from:"Tristan Croll"

Re: [ccp4bb] Odd Positive Density Around a Cystine

2022-08-10 Thread Tristan Croll

I've seen this happen with B-factor refinement reasonably often. As I 
understand it the basic problem is that for small B-factors the gradient 
d(density)d(B) is large, but for large B-factors the gradient is small. So if 
the starting B-factor on an atom is very low and substantially lower than what 
the density supports, then the first refinement cycle can overstep to the point 
where there's almost no gradient, so future refinement steps don't pull it back 
down. Tightening the B-factor similarity restraints to surrounding atoms can 
help.

From: CCP4 bulletin board  on behalf of Thomas, Leonard 
M. 
Sent: 10 August 2022 17:01
To: CCP4BB@JISCMAIL.AC.UK 
Subject: Re: [ccp4bb] Odd Positive Density Around a Cystine

While the B value in the starting model was fine.  I did drop it to a 
reasonable value and it now seems to be behaving, was thinking about trying 
that before I posted.  Since this was an  MR determination I did check and the 
starting model B value was actually smaller then the surrounding residues.  I 
guess some thing went odd with the first round of refinement.

Thank you for the suggestions.

Len


> On Aug 10, 2022, at 10:25 AM, Dale Tronrud  wrote:
>
>
>   A large B value with positive difference density sure implies a convergence 
> problem with the refinement.  Was the B value extreme in your starting model? 
>  (A starting B that is wildly too large or too small at the start may cause 
> it to become trapped in the refinement.) Maybe if you rerun your refinement 
> with a moderate starting value for the B you will end up a more sensible 
> result.
>
>   The only other way to end up with a parameter that directly conflicts with 
> the difference density is a bad restraint, but that doesn't sound likely 
> based on your description.
>
> Dale Tronrud
>
> On 8/10/2022 7:59 AM, Thomas, Leonard M. wrote:
>> Hello All,
>> I have run into something odd.  In working on a structure for one of the 
>> groups I work with regularly, on one of the cystine residues I have a very 
>> large positive density peak at the sulfur position. The B value is 
>> approximately 4 times the other values in the residue and on other cystine 
>> residues.  The overall structure has 2 molecules in the asymmetric unit  and 
>> the corresponding cystine  on the other monomer is behaving as I would 
>> expect.   There are no disulfides in the structure.
>> The data were collected on 9-2 at SSRL and all three of the data sets we 
>> collected show the same thing, all data go to about 2.2 angstroms.  We are 
>> trying to determine the ligand binding in the molecule but this cystine is 
>> not involved in ligand binding.  In house and other synchrotron data from 
>> previous protein preps and data collection runs of the same molecule grown 
>> in very similar condition and crystallized in the same space group have the 
>> residue behaving normally.
>> I am open to any ideas as to what may be going on as I am rather puzzled by 
>> this.
>> Thanks for any input,
>> Len Thomas
>> Leonard Thomas, Ph.D.
>> Biomolecular Structure Core, Director
>> Oklahoma COBRE in Structural Biology
>> Price Family Foundation Institute of Structural Biology
>> University of Oklahoma
>> Department of Chemistry and Biochemistry
>> 101 Stephenson Parkway
>> Norman, OK 73019-5251
>> Office: (405)325-1126
>> lmtho...@ou.edu 
>> https://eur03.safelinks.protection.outlook.com/?url=http%3A%2F%2Fwww.ou.edu%2Fstructuralbiology%2Fcobre-core-facilities%2Fmcldata=05%7C01%7Ctic20%40universityofcambridgecloud.onmicrosoft.com%7C993e277f8ad542f6a9ae08da7ae9bba0%7C49a50445bdfa4b79ade3547b4f3986e9%7C0%7C0%7C637957441549582999%7CUnknown%7CTWFpbGZsb3d8eyJWIjoiMC4wLjAwMDAiLCJQIjoiV2luMzIiLCJBTiI6Ik1haWwiLCJXVCI6Mn0%3D%7C3000%7C%7C%7Csdata=chxTRCdc%2FODmxM7NzVaZcAaXDJ0zMDulcWQrF%2FERs7A%3Dreserved=0
>>  
>> 
>> 
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>>  
>>

Re: [ccp4bb] Validation of structure prediction

2022-01-13 Thread Tristan Croll

Hard but not impossible - even when you *are* fitting to low-res density. See
https://twitter.com/crolltristan/status/1381258326223290373?s=21 for example -
no Ramachandran outliers, 1.3% sidechain outliers, clashscore of 2... yet
multiple regions out of register by anywhere up to 15 residues! I never
publicly named the structure (although I did share my rebuilt model with the
authors), but the videos and images in that thread should be enough to
illustrate the scale of the problem.

And that was *with* a map to fit! Take away the map, and run some MD energy
minimisation (perhaps with added Ramachandran and rotamer restraints), and I
think it would be easy to get your model to fool most “simple” validation
metrics (please don’t actually do this). The upshot is that I still think
validation of predicted models in the absence of at least moderate-resolution
experimental data is still a major challenge requiring very careful thought.

— Tristan

On 13 Jan 2022, at 18:41, James Holton
mailto:jmhol...@lbl.gov>> wrote:

Agree with Pavel.

Something I think worth adding is a reminder that the MolProbity score only
looks at bad clashes, ramachandran and rotamer outliers.

MPscore=0.426∗ln(1+clashscore)+0.33∗ln(1+max(0,rota_out−1))+0.25∗ln(1+max(0,rama_iffy−2))+0.5

It pays no attention whatsoever to twisted peptide bonds, C-beta deviations,
and, for that matter, bond lengths and bond angles. If you tweak your weights
right you can get excellent MP scores, but horrible "geometry" in the
traditional bonds-and-angles sense. The logic behind this kind of validation is
that normally nonbonds and torsions are much softer than bond and angle
restraints and therefore fertile ground for detecting problems. Thus far, I am
not aware of any "Grand Unified Score" that combines all geometric
considerations, but perhaps it is time for one?

Tristan's trivial solution aside, it is actually very hard to make all the
"geometry" ideal for a real-world fold, and especially difficult to do without
also screwing up the agreement with density (R factor). I would argue that if
you don't have an R factor then you should get one, but I am interested in
opinions about alternatives.

I.E. What if we could train an AI to predict Rfree by looking at the
coordinates?

-James Holton
MAD Scientist

On 12/21/2021 9:25 AM, Pavel Afonine wrote:
Hi Reza,
If you think about it this way... Validation is making sure that the model
makes sense, data make sense and model-to-data fit make sense, then the answer
to your question is obvious: in your case you do not have experimental data (at
least in a way we used to think of it) and so then of these three validation
items you only have one, which, for example, means you don’t have to report
things like R-factors or completeness in high-resolution shell.
Really, the geometry of an alpha helix does not depend on how you determined
it: using X-rays or cryo-EM or something else! So, most (if not all) model
validation tools still apply.
Pavel

On Mon, Dec 20, 2021 at 8:10 AM Reza Khayat
mailto:rkha...@ccny.cuny.edu>> wrote:

Hi,

Can anyone suggest how to validate a predicted structure? Something similar to
wwPDB validation without the need for refinement statistics. I realize this is
a strange question given that the geometry of the model is anticipated to be
fine if the structure was predicted by a server that minimizes the geometry to
improve its statistics. Nonetheless, the journal has asked me for such a
report. Thanks.

Best wishes,

Reza

Reza Khayat, PhD
Associate Professor
City College of New York
Department of Chemistry and Biochemistry
New York, NY 10031

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Re: [ccp4bb] Validation of structure prediction

2021-12-21 Thread Tristan Croll

I agree with Dale. Tools like MolProbity are not the right approach to 
validating a structure prediction. To understand why, just consider that all 
you need to do to get a perfect MolProbity score is predict every structure as 
a single long alpha helix with ideal rotamers, with a kink at each proline. 

To validate a predicted structure will require a completely different toolset - 
one that I’m not sure fully exists yet. 

— Tristan 

> On 20 Dec 2021, at 18:47, Dale Tronrud  wrote:
> 
>   I don't see any reason to believe that software designed to validate 
> crystallographic or NMR models would have any utility validating AlphaFold 
> predicted models.  Doesn't the prediction software already ensure that all 
> the indicators used by Molprobity are obeyed?  I'm afraid that the tools to 
> validate any new technique must be designed specifically for that technique. 
> (And when they become available they will be useless for validating 
> crystallographic models!)
> 
> Dale E. Tronrud
> 
>> On 12/20/2021 10:28 AM, Nicholas Clark wrote:
>> The Molprobity server can be run online and only requires the coordinates in 
>> PDB format: http://molprobity.biochem.duke.edu/ 
>> .
>> Best,
>> Nick Clark
>> On Mon, Dec 20, 2021 at 11:10 AM Reza Khayat > > wrote:
>>Hi,
>>Can anyone suggest how to validate a predicted structure? Something
>>similar to wwPDB validation without the need for refinement
>>statistics. I realize this is a strange question given that the
>>geometry of the model is anticipated to be fine if the structure was
>>predicted by a server that minimizes the geometry to improve its
>>statistics. Nonetheless, the journal has asked me for such a report.
>>Thanks.
>>Best wishes,
>>Reza
>>Reza Khayat, PhD
>>Associate Professor
>>City College of New York
>>Department of Chemistry and Biochemistry
>>New York, NY 10031
>>
>>To unsubscribe from the CCP4BB list, click the following link:
>>https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1
>>
>> -- 
>> Nicholas D. Clark
>> PhD Candidate
>> Malkowski Lab
>> University at Buffalo
>> Department of Structural Biology
>> Jacob's School of Medicine & Biomedical Sciences
>> 955 Main Street, RM 5130
>> Buffalo, NY 14203
>> Cell: 716-830-1908
>> 
>> To unsubscribe from the CCP4BB list, click the following link:
>> https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1 
>> 
> 
> 
> 
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Re: [ccp4bb] saving coordinates with respect to map in ChimeraX

2021-06-28 Thread Tristan Croll

The way things generally work in ChimeraX is that each model/map is stored 
internally with its original coordinates unchanged, along with one or more 
transforms placing it into scene coordinates. Saving model A relative to 
model/map B means that the coordinates to be saved are transformed such that A 
overlays with the original B in the same way you see in the scene. That can get 
a little complex if both model and map have been moved... if you just want to 
save things exactly as they are shown, uncheck the checkbox entirely and 
everything will be written in scene coordinates.

-- Tristan

From: Jon Cooper 
Sent: 28 June 2021 13:43
To: Tristan Croll ; CCP4BB@JISCMAIL.AC.UK 

Subject: Re: [ccp4bb] saving coordinates with respect to map in ChimeraX

It's not important, but I am curious what 'Save relative to model/map' actually 
means in that widget. Is it just a case of copying the relevant CRYST1 card? I 
think Faisal is asking how to save new/modified coordinates but probably I 
misunderstood...

Cheers, Jon.C.


Sent from ProtonMail mobile



 Original Message 
On 28 Jun 2021, 10:32, Tristan Croll < ti...@cam.ac.uk> wrote:

Hi Faisal,

Are you using the latest version (1.2.5)? That should give you a dialog that 
looks something like this.

Best regards,
Tristan


[cid:c60bb51b-6f11-4646-9800-6ce976abc3f0]

From: khaja faisal tarique 
Sent: 28 June 2021 10:29
To: Tristan Croll 
Cc: CCP4BB@JISCMAIL.AC.UK 
Subject: Re: [ccp4bb] saving coordinates with respect to map in ChimeraX

Hi Tristan

I have posted my question in ChimeraX mailing list as well and waiting for 
their reply.

What you are suggesting is for Chimera. I dont see any such option in ChimeraX. 
That save relative to section does not appear here or may be there is a 
different way of saving model relative to map in ChimeraX.

Please correct me if I am wrong.

Faisal

On Mon, 28 Jun 2021, 2:44 pm Tristan Croll, 
mailto:ti...@cam.ac.uk>> wrote:
Hi Faisal,

This is really a question for the ChimeraX-users list 
(chimerax-us...@cgl.ucsf.edu<mailto:chimerax-us...@cgl.ucsf.edu>). But the 
quick run-down: using the File/Save dialogue, you'll see a checkbox with "Save 
relative to model: " and then a drop-down menu. Just choose the map in the 
drop-down menu, and you should get the result you want.

Best regards,
Tristan

From: CCP4 bulletin board mailto:CCP4BB@JISCMAIL.AC.UK>> 
on behalf of khaja faisal tarique 
mailto:khajafaisaltari...@gmail.com>>
Sent: 28 June 2021 09:43
To: CCP4BB@JISCMAIL.AC.UK<mailto:CCP4BB@JISCMAIL.AC.UK> 
mailto:CCP4BB@JISCMAIL.AC.UK>>
Subject: [ccp4bb] saving coordinates with respect to map in ChimeraX

Hello everyone

While doing rigid body fit we know that after placing the coordinate to its 
correct position we can save it with respect to the map. Does anyone know if a 
similar feature exists in ChimeraX ?  Is there a way where the coordinates 
.pdb/cif file can be saved with respect to its map in ChimeraX ?

Thank you

--
Faisal Tarique Khaja



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Re: [ccp4bb] saving coordinates with respect to map in ChimeraX

2021-06-28 Thread Tristan Croll

Hi Faisal,

Are you using the latest version (1.2.5)? That should give you a dialog that 
looks something like this.

Best regards,
Tristan


[cid:c60bb51b-6f11-4646-9800-6ce976abc3f0]

From: khaja faisal tarique 
Sent: 28 June 2021 10:29
To: Tristan Croll 
Cc: CCP4BB@JISCMAIL.AC.UK 
Subject: Re: [ccp4bb] saving coordinates with respect to map in ChimeraX

Hi Tristan

I have posted my question in ChimeraX mailing list as well and waiting for 
their reply.

What you are suggesting is for Chimera. I dont see any such option in ChimeraX. 
That save relative to section does not appear here or may be there is a 
different way of saving model relative to map in ChimeraX.

Please correct me if I am wrong.

Faisal

On Mon, 28 Jun 2021, 2:44 pm Tristan Croll, 
mailto:ti...@cam.ac.uk>> wrote:
Hi Faisal,

This is really a question for the ChimeraX-users list 
(chimerax-us...@cgl.ucsf.edu<mailto:chimerax-us...@cgl.ucsf.edu>). But the 
quick run-down: using the File/Save dialogue, you'll see a checkbox with "Save 
relative to model: " and then a drop-down menu. Just choose the map in the 
drop-down menu, and you should get the result you want.

Best regards,
Tristan

From: CCP4 bulletin board mailto:CCP4BB@JISCMAIL.AC.UK>> 
on behalf of khaja faisal tarique 
mailto:khajafaisaltari...@gmail.com>>
Sent: 28 June 2021 09:43
To: CCP4BB@JISCMAIL.AC.UK<mailto:CCP4BB@JISCMAIL.AC.UK> 
mailto:CCP4BB@JISCMAIL.AC.UK>>
Subject: [ccp4bb] saving coordinates with respect to map in ChimeraX

Hello everyone

While doing rigid body fit we know that after placing the coordinate to its 
correct position we can save it with respect to the map. Does anyone know if a 
similar feature exists in ChimeraX ?  Is there a way where the coordinates 
.pdb/cif file can be saved with respect to its map in ChimeraX ?

Thank you

--
Faisal Tarique Khaja



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Re: [ccp4bb] saving coordinates with respect to map in ChimeraX

2021-06-28 Thread Tristan Croll

Hi Faisal,

This is really a question for the ChimeraX-users list 
(chimerax-us...@cgl.ucsf.edu). But the quick run-down: using the File/Save 
dialogue, you'll see a checkbox with "Save relative to model: " and then a 
drop-down menu. Just choose the map in the drop-down menu, and you should get 
the result you want.

Best regards,
Tristan

From: CCP4 bulletin board  on behalf of khaja faisal 
tarique 
Sent: 28 June 2021 09:43
To: CCP4BB@JISCMAIL.AC.UK 
Subject: [ccp4bb] saving coordinates with respect to map in ChimeraX

Hello everyone

While doing rigid body fit we know that after placing the coordinate to its 
correct position we can save it with respect to the map. Does anyone know if a 
similar feature exists in ChimeraX ?  Is there a way where the coordinates 
.pdb/cif file can be saved with respect to its map in ChimeraX ?

Thank you

--
Faisal Tarique Khaja



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Re: [ccp4bb] Enzyme Vmax and Km

2021-06-18 Thread Tristan Croll

Hi Prem,

The immediate problem here is that the curve for the processed substrate simply 
cannot be described by simple Michaelis-Menten kinetics. Assuming the assay has 
worked as expected, the declining rate with increasing substrate concentration 
suggests to me that this substrate also acts as an allosteric inhibitor, so 
assays at high [substrate] will make it *look* like the unprocessed substrate 
is preferred even though the processed one is cleaved faster by the uninhibited 
enzyme.

Hope this helps,
Tristan

On 18 Jun 2021, at 05:05, Prem Prakash 
mailto:prem...@gmail.com>> wrote:

Dear all,
Sorry for this off topic. I am working on an enzyme that has an exonuclease 
activity. The enzyme preferentially cleaves an unprocessed substrate at a 
faster rate than the processed one (known by qualitative analysis). Recently, I 
calculated the Vmax, Km and kcat of the enzyme for unprocessed substrate which 
are 18.2 pmol/min, 182 nM and 7.1 sec-1 respectively. However, the Processed 
substrate has apparently a lower range of Km (not calculated) as reflected from 
the curve (because the same increasing concentration range which is used for 
unprocessed, shows a steep decline in the initial velocity of the enzyme with 
processed substrate.
The latter suggests that Km is way lower than expected. In this case, the 
question is, if the Km of processed substrate is way lower than the 
Unprocessed, how can we see a faster rate with the former substrate than later. 
i.e lower Km and slower rate of cleavage. If it's possible please give some 
insights. I have attached the plot comparison between two kinetic assays.

With kind regards,

Prem





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Re: [ccp4bb] Analysis of NMR ensembles

2021-05-26 Thread Tristan Croll

(sending properly this time, rather than just to Rasmus)

I don't believe a "color by RMS" option is in ChimeraX right now (I'll suggest 
it to the developers), but this will align all models then set B-factors for 
each residue to the RMS CA deviation from the mean position. Could be extended 
fairly trivially to do all-atom RMS if you wanted to. Change the extension for 
the attached text file to .py, open your NMR ensemble in ChimeraX, select all 
the models (typically just "sel #1" should do the trick), then use File/Open 
and choose color_by_rms.py (or just "open color_by_rms.py" on the command line 
if it's in your working directory). As long as all models have the same set of 
residues, it should do the trick. Example image for 2kv5 attached.

Have fun!

-- Tristan

From: CCP4 bulletin board  on behalf of Harry Powell - 
CCP4BB <193323b1e616-dmarc-requ...@jiscmail.ac.uk>
Sent: 26 May 2021 16:04
To: CCP4BB@JISCMAIL.AC.UK 
Subject: [ccp4bb] Analysis of NMR ensembles

Hi

Given that there are plenty of people on this BB who are structural biologists 
rather than “just” crystallographers, I thought someone here might be able to 
help.

If I have a structure in the PDB (e.g. 2kv5) that is an ensemble of structures 
that fit the NOEs, is there a tool available that will give me some idea about 
the bits of the structure that do not vary much (“rigid”) and the bits that are 
all over the place (“flexible”)?

Would superpose or gesamt be a good tool for this? Ideally I’d like something 
that could add a figure to the B columns in a PDB file so I could see something 
in QTMG (or PyMol if forced…) or do other useful things with the information.

Harry


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from chimerax.atomic import Residues, Atoms, selected_residues
from math import sqrt
import numpy
models = selected_residues(session).unique_structures
from chimerax.core.commands import run

run(session, f'match {"|".join(["#"+m.id_string for m in models[1:]])} to 
#{models[0].id_string}')

for residues in zip(*[m.residues for m in models]):
residues = Residues(residues)
pas = Atoms(residues.principal_atoms)
if not len(pas):
residues.atoms.bfactors = 0
continue

coords = pas.scene_coords
avg = numpy.mean(coords, axis=0)
deviations = coords-avg
distances = numpy.linalg.norm(deviations, axis=0)
rms = sqrt(numpy.mean(distances**2))
residues.atoms.bfactors = rms

run(session, f'color bfactor sel')



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[ccp4bb] Release of ISOLDE 1.2

2021-05-13 Thread Tristan Croll

Hi all,

I'm very happy to announce the release of ISOLDE 1.2, now available for 
installation into the ChimeraX 1.2 release. To get it, first download and 
install ChimeraX 1.2 from https://www.rbvi.ucsf.edu/chimerax/download.html, 
then go to "Tools/More Tools..." in the menu and follow the links to install 
ISOLDE. Key new features include:


  *   The ability to add residues to the ends of protein chains (see 
https://twitter.com/CrollTristan/status/1391747032579289089?s=20; support for 
nucleic acid chain extension still in development)
  *   Lower-fidelity simulation modes for greatly improved performance on 
lower-end GPUs (see 
https://twitter.com/CrollTristan/status/1390709520876331009?s=20)
  *   Commands to write input files for phenix.refine and 
phenix.real_space_refine with recommended settings for refining models from 
ISOLDE
  *   Command to write a ProSMART-like restraint definition file to softly 
restrain the model to its current geometry in Refmac
  *   Improvements to stability and performance of bulk solvent/scaling in 
crystallographic datasets
  *   Short aliases for commonly-used commands to make life easier for power 
users.

Please enjoy!

Best regards,
Tristan




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Re: [ccp4bb] Bug in mmCIF handling of UNK residues?

2021-02-13 Thread Tristan Croll

Browsing backwards through a dozen or so of the most recent UNK-containing 
structures, I haven't found a counter-example yet - apart from those where the 
UNK residues are a single contiguous stretch and given their own chain ID. So a 
recent problem?

From: Philip D. Jeffrey 
Sent: 12 February 2021 19:56
To: CCP4BB@JISCMAIL.AC.UK ; Tristan Croll 

Subject: Re: Bug in mmCIF handling of UNK residues?

Doesn't seem to be the case for all instances: that table isn't present in 5BV0 
despite the N-terminal residues of Nyv1 being modeled as UNK in the 
Vps16:Vps33:Nyv1 complex due to a symmetry overlap.  Nyv1, C165-179 are UNK 
with partial occupancy, which is the N-terminal part of the model for that 
chain, then there's a gap of 3 missing residues, and then there's polypeptide 
model which we've assigned to sequence, all in the same chain.

Not sure if the missing table is something that turned up after the deposition 
date (June 2015), or if it's related to the missing residues between the UNK 
segment and the defined amino acids.

Phil Jeffrey
Princeton

From: CCP4 bulletin board  on behalf of Tristan Croll 

Sent: Friday, February 12, 2021 1:03 PM
To: CCP4BB@JISCMAIL.AC.UK 
Subject: [ccp4bb] Bug in mmCIF handling of UNK residues?

Hi all,

If I open (as far as I can tell) the mmCIF for any structure in the wwPDB that 
contains both defined amino acids and UNK in the same chain, then the UNK 
section is treated as covalently bonded to the flanking sequence. This appears 
to be a bug in the mmCIF generation itself, not in the viewing software 
(ChimeraX, in this case): if I look in 7kzn as a random example, I see:

loop_
_pdbx_validate_polymer_linkage.id
_pdbx_validate_polymer_linkage.PDB_model_num
_pdbx_validate_polymer_linkage.auth_atom_id_1
_pdbx_validate_polymer_linkage.auth_asym_id_1
_pdbx_validate_polymer_linkage.auth_comp_id_1
_pdbx_validate_polymer_linkage.auth_seq_id_1
_pdbx_validate_polymer_linkage.PDB_ins_code_1
_pdbx_validate_polymer_linkage.label_alt_id_1
_pdbx_validate_polymer_linkage.auth_atom_id_2
_pdbx_validate_polymer_linkage.auth_asym_id_2
_pdbx_validate_polymer_linkage.auth_comp_id_2
_pdbx_validate_polymer_linkage.auth_seq_id_2
_pdbx_validate_polymer_linkage.PDB_ins_code_2
_pdbx_validate_polymer_linkage.label_alt_id_2
_pdbx_validate_polymer_linkage.dist
1 1 C X UNK 345 ? ? N X UNK 348 ? ? 10.08
2 1 C X UNK 396 ? ? N X UNK 403 ? ? 28.65
3 1 C Y UNK 281 ? ? N Y UNK 284 ? ? 6.72
4 1 C Y UNK 387 ? ? N Y UNK 394 ? ? 22.26
5 1 C Y UNK 420 ? ? N Y UNK 424 ? ? 12.82

Considering that the coords themselves generally seem fine, I guess this is 
happening post deposition?

Best regards,

Tristan

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[ccp4bb] Bug in mmCIF handling of UNK residues?

2021-02-12 Thread Tristan Croll

Hi all,

If I open (as far as I can tell) the mmCIF for any structure in the wwPDB that 
contains both defined amino acids and UNK in the same chain, then the UNK 
section is treated as covalently bonded to the flanking sequence. This appears 
to be a bug in the mmCIF generation itself, not in the viewing software 
(ChimeraX, in this case): if I look in 7kzn as a random example, I see:


loop_
_pdbx_validate_polymer_linkage.id
_pdbx_validate_polymer_linkage.PDB_model_num
_pdbx_validate_polymer_linkage.auth_atom_id_1
_pdbx_validate_polymer_linkage.auth_asym_id_1
_pdbx_validate_polymer_linkage.auth_comp_id_1
_pdbx_validate_polymer_linkage.auth_seq_id_1
_pdbx_validate_polymer_linkage.PDB_ins_code_1
_pdbx_validate_polymer_linkage.label_alt_id_1
_pdbx_validate_polymer_linkage.auth_atom_id_2
_pdbx_validate_polymer_linkage.auth_asym_id_2
_pdbx_validate_polymer_linkage.auth_comp_id_2
_pdbx_validate_polymer_linkage.auth_seq_id_2
_pdbx_validate_polymer_linkage.PDB_ins_code_2
_pdbx_validate_polymer_linkage.label_alt_id_2
_pdbx_validate_polymer_linkage.dist
1 1 C X UNK 345 ? ? N X UNK 348 ? ? 10.08
2 1 C X UNK 396 ? ? N X UNK 403 ? ? 28.65
3 1 C Y UNK 281 ? ? N Y UNK 284 ? ? 6.72
4 1 C Y UNK 387 ? ? N Y UNK 394 ? ? 22.26
5 1 C Y UNK 420 ? ? N Y UNK 424 ? ? 12.82


Considering that the coords themselves generally seem fine, I guess this is 
happening post deposition?

Best regards,

Tristan



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Re: [ccp4bb] ideal ligands

2021-02-06 Thread Tristan Croll

I know . Just suggesting the most likely cause of the problem.

-- T

From: Bernhard Rupp 
Sent: 06 February 2021 14:38
To: Tristan Croll 
Cc: CCP4BB@jiscmail.ac.uk 
Subject: Re: [ccp4bb] ideal ligands

That is what 'uranium atom solution' implies:
All atoms collapsing into one :-)

Cheers br

On Sat, Feb 6, 2021, 06:34 Tristan Croll 
mailto:ti...@cam.ac.uk>> wrote:
The ideal coordinates in the CCD entries for BCR and LUT are null - my guess is 
that in these cases all the coordinates just default to (0,0,0) in the PDBeChem 
script.

From: CCP4 bulletin board mailto:CCP4BB@JISCMAIL.AC.UK>> 
on behalf of Bernhard Rupp 
mailto:hofkristall...@gmail.com>>
Sent: 05 February 2021 20:51
To: CCP4BB@JISCMAIL.AC.UK<mailto:CCP4BB@JISCMAIL.AC.UK> 
mailto:CCP4BB@JISCMAIL.AC.UK>>
Subject: [ccp4bb] ideal ligands


Hi Fellows,



PDBeChem under ‘ideal’ coordinates returns a small molecule uranium atom 
solution for BCR, LUT, and possibly other xanthophylls, eg.

ftp://ftp.ebi.ac.uk/pub/databases/msd/pdbechem_v2/B/BCR/BCR_ideal.pdb

while the representative files are ok.



Is this intended?



Thx, BR

--

Bernhard Rupp

http://www.hofkristallamt.org/

b...@hofkristallamt.org<mailto:b...@hofkristallamt.org>

+1 925 209 7429

+43 676 571 0536

--

Many plausible ideas vanish

at the presence of thought

--





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Re: [ccp4bb] ideal ligands

2021-02-06 Thread Tristan Croll

The ideal coordinates in the CCD entries for BCR and LUT are null - my guess is 
that in these cases all the coordinates just default to (0,0,0) in the PDBeChem 
script.

From: CCP4 bulletin board  on behalf of Bernhard Rupp 

Sent: 05 February 2021 20:51
To: CCP4BB@JISCMAIL.AC.UK 
Subject: [ccp4bb] ideal ligands


Hi Fellows,



PDBeChem under ‘ideal’ coordinates returns a small molecule uranium atom 
solution for BCR, LUT, and possibly other xanthophylls, eg.

ftp://ftp.ebi.ac.uk/pub/databases/msd/pdbechem_v2/B/BCR/BCR_ideal.pdb

while the representative files are ok.



Is this intended?



Thx, BR

--

Bernhard Rupp

http://www.hofkristallamt.org/

b...@hofkristallamt.org

+1 925 209 7429

+43 676 571 0536

--

Many plausible ideas vanish

at the presence of thought

--





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Re: [ccp4bb] External: Re: [ccp4bb] AlphaFold: more thinking and less pipetting (?)

2020-12-11 Thread Tristan Croll

I'm not Randy, but I do have an answer: like this. This is T1049-D1. AlphaFold 
prediction in red, experimental structure (6y4f) in green. Agreement is close 
to perfect, apart from the C-terminal tail which is way off - but clearly 
flexible and only resolved in this conformation in the crystal due to packing 
interactions. GDT_TS is 93.1; RMS_CA is 3.68 - but if you exclude those tail 
residues, it's 0.79. With an alignment cutoff of 1 A, you can align 109 of 134 
CAs with an RMSD of 0.46 A.

From: CCP4 bulletin board  on behalf of Leonid Sazanov 

Sent: 11 December 2020 10:36
To: CCP4BB@JISCMAIL.AC.UK 
Subject: Re: [ccp4bb] External: Re: [ccp4bb] AlphaFold: more thinking and less 
pipetting (?)

Dear Randy,

Can you comment on why for some of AplhaFold2 models with GDT_TS > 90 
(supposedly as good as experimental model) the RMS_CA (backbone) is > 3.0 
Angstrom? Such a deviation can hardly be described as good as experimental. 
Could it be that GDT_TS is kind of designed to evaluate how well the general 
sub-domain level fold is predicted, rather than overall detail?

Thanks,
Leonid


>
Several people have mentioned lack of peer review as a reason to doubt the 
significance of the AlphaFold2 results.  There are different routes to peer 
review and, while the results have not been published in a peer review journal, 
I would have to say (as someone who has been an assessor for two CASPs, as well 
as having editorial responsibilities for a peer-reviewed journal), the peer 
review at CASP is much more rigorous than the peer review that most papers 
undergo.  The targets are selected from structures that have recently been 
solved but not published or disseminated, and even just tweeting a C-alpha 
trace is probably enough to get a target cancelled.  In some cases (as we’ve 
heard here) the people determining the structure are overly optimistic about 
when their structure solution will be finished, so even they may not know the 
structure at the time it is predicted.  The assessors are blinded to the 
identities of the predictors, and they carry out months of calculations and 
inspections of the models, computing ranking scores before they find out who 
made the predictions.  Most assessors try to bring something new to the 
assessment, because the criteria should get more stringent as the predictions 
get better, and they have new ideas of what to look for, but there’s always 
some overlap with “traditional” measures such as GDT-TS, GDT-HA (more stringent 
high-accuracy version of GDT) and lDDT.



Of course we’d all like to know the details of how AlphaFold2 works, and the 
DeepMind people could have been (and should be) much more forthcoming, but 
their results are real.  They didn’t have any way of cheating, being selective 
about what they reported, or gaming the system in any other way that the other 
groups couldn’t do.  (And yes, when we learned that DeepMind was behind the 
exceptionally good results two years ago at CASP13, we made the same half-jokes 
about whether Gmail had been in the database they were mining!)



Best wishes,



Randy Read



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Re: [ccp4bb] External: Re: [ccp4bb] AlphaFold: more thinking and less pipetting (?)

2020-12-08 Thread Tristan Croll

... and of course I meant "between model and target".
____
From: Tristan Croll 
Sent: 08 December 2020 16:35
To: CCP4BB@JISCMAIL.AC.UK ; Marko Hyvonen 

Subject: Re: [ccp4bb] External: Re: [ccp4bb] AlphaFold: more thinking and less 
pipetting (?)

An example: this is TS1038-D1 - designated by the CASP organisers as in the 
"free modelling" category due to the absence of any close homologues in the 
wwPDB. The experimental model is in tan, the AlphaFold2 prediction in cyan. As 
far as I'm concerned, the only way to describe this is "nailed it". Using 
ChimeraX's MatchMaker to do the alignment, 84 of 114 residues align to a 
CA-RMSD of 0.57 A, (2.3 A across all residues, with the outliers being one 
flexible-looking loop and the N-terminal tail). Further than that, it's nailed 
almost all the details - if you exclude surface-exposed residues, I count less 
than half a dozen sidechains with significantly different rotamers compared to 
the template. The upshot is that the difference between model and template 
appears easily within the range of variation you'd expect to see between 
different crystal forms of the same protein.

For comparison, the next best group got the three-strand beta-sheet at bottom 
right essentially correct, but everything else (apart from the vague fold) 
wrong. MatchMaker aligns 28 CA atoms with an RMSD of 0.64 A, but the overall 
CA-RMSD blows out to 9.6 A. So I don't think there's any denying that this is a 
spectacular advance that will change the field markedly.

Best regards,

Tristan

From: CCP4 bulletin board  on behalf of Marko Hyvonen 

Sent: 08 December 2020 15:07
To: CCP4BB@JISCMAIL.AC.UK 
Subject: Re: [ccp4bb] External: Re: [ccp4bb] AlphaFold: more thinking and less 
pipetting (?)

Hi Ian,

The data on Alphafold2 target RMSDs seems to be correct, but that "resolution 
around 2.5Å", makes no sense, I agree  - had not noticed that before. I can see 
that this has been raised in the Twitter feed comments to his post too.

I was highlighting this more for the alternative viewpoint on the discussion 
and also on the interesting detail on the resources needed/available (assuming 
correct!).

Marko

On 08/12/2020 14:02, Ian Tickle wrote:

Hi Marko

I hope he hasn't confused resolution with RMSD error:

"Just keep in mind that (1) a lower RMSD represents a better predicted 
structure, and that (2) most experimental structures have a resolution around 
2.5 Å. Taking this into consideration, about a third (36%) of Group 427’s 
submitted targets were predicted with a root-mean-square deviation (RMSD) under 
2 Å, and 86% were under 5 Å, with a total mean of 3.8 Å."

Cheers

-- Ian

On Tue, 8 Dec 2020 at 13:51, Marko Hyvonen 
mailto:mh...@cam.ac.uk>> wrote:
Here is another take on this topic, by Carlos Quteiral (@c_outeiral), from a 
non-crystallographer's point of view, covering many of the points discussed in 
this thread  (incl. an example of the model guiding correction of the 
experimental structure).

https://www.blopig.com/blog/2020/12/casp14-what-google-deepminds-alphafold-2-really-achieved-and-what-it-means-for-protein-folding-biology-and-bioinformatics/

Marko

On 08/12/2020 13:25, Tristan Croll wrote:
This is a number that needs to be interpreted with some care. 2 Å crystal 
structures in general achieve an RMSD of 0.2 Å on the portion of the crystal 
that's resolved, including loops that are often only in well-resolved 
conformations due to physiologically-irrelevant crystal packing interactions. 
The predicted models, on the other hand, are in isolation. Once you get to the 
level achieved by this last round of predictions, that starts making fair 
comparison somewhat more difficult*. Two obvious options that I see: (1) limit 
the comparison only to the stable core of the protein (in which case many of 
the predictions have RMSDs in the very low fractions of an Angstrom), or (2) 
compare ensembles derived from MD simulations starting from the experimental 
and predicted structure, and see how well they overlap.

-- Tristan

* There's one more thorny issue when you get to this level: it becomes more and 
more possible (even likely) that the prediction gets some things right that are 
wrong in the experimental structure.

From: CCP4 bulletin board <mailto:CCP4BB@JISCMAIL.AC.UK> 
on behalf of Ian Tickle <mailto:ianj...@gmail.com>
Sent: 08 December 2020 13:04
To: CCP4BB@JISCMAIL.AC.UK<mailto:CCP4BB@JISCMAIL.AC.UK> 
<mailto:CCP4BB@JISCMAIL.AC.UK>
Subject: Re: [ccp4bb] External: Re: [ccp4bb] AlphaFold: more thinking and less 
pipetting (?)

There was a little bit of press-release hype: the release stated "a score of 
around 90 GDT is informally considered to be competitive with results obtained 
from experimental methods" and "our latest AlphaFold system achieves a median 
score of 92.

Re: [ccp4bb] External: Re: [ccp4bb] AlphaFold: more thinking and less pipetting (?)

2020-12-08 Thread Tristan Croll

This is a number that needs to be interpreted with some care. 2 Å crystal 
structures in general achieve an RMSD of 0.2 Å on the portion of the crystal 
that's resolved, including loops that are often only in well-resolved 
conformations due to physiologically-irrelevant crystal packing interactions. 
The predicted models, on the other hand, are in isolation. Once you get to the 
level achieved by this last round of predictions, that starts making fair 
comparison somewhat more difficult*. Two obvious options that I see: (1) limit 
the comparison only to the stable core of the protein (in which case many of 
the predictions have RMSDs in the very low fractions of an Angstrom), or (2) 
compare ensembles derived from MD simulations starting from the experimental 
and predicted structure, and see how well they overlap.

-- Tristan

* There's one more thorny issue when you get to this level: it becomes more and 
more possible (even likely) that the prediction gets some things right that are 
wrong in the experimental structure.

From: CCP4 bulletin board  on behalf of Ian Tickle 

Sent: 08 December 2020 13:04
To: CCP4BB@JISCMAIL.AC.UK 
Subject: Re: [ccp4bb] External: Re: [ccp4bb] AlphaFold: more thinking and less 
pipetting (?)

There was a little bit of press-release hype: the release stated "a score of 
around 90 GDT is informally considered to be competitive with results obtained 
from experimental methods" and "our latest AlphaFold system achieves a median 
score of 92.4 GDT overall across all targets. This means that our predictions 
have an average error 
(RMSD)
 of approximately 1.6 Angstroms,".

Experimental methods achieve an average error of around 0.2 Ang. or better at 2 
Ang. resolution, and of course much better at atomic resolution (1 Ang. or 
better), or around 0.5 Ang. at 3 Ang. resolution.  For ligand-binding studies I 
would say you need 3 Ang. resolution or better.  1.6 Ang. error is probably 
equivalent to around 4 Ang. resolution.  No doubt that will improve with time 
and experience, though I think it will be an uphill struggle to get better than 
1 Ang. error, simply because the method can't be better than the data that go 
into it and 1-1.5 Ang. represents a typical spread of homologous models in the 
PDB.  So yes very competitive if you're desperate for a MR starting model, but 
not quite yet there for a refined high-resolution structure.

Cheers

-- Ian

On Tue, 8 Dec 2020 at 12:11, Harry Powell - CCP4BB 
<193323b1e616-dmarc-requ...@jiscmail.ac.uk>
 wrote:
Hi

It’s a bit more than science by press release - they took part in CASP14 where 
they were given sequences but no other experimental data, and did significantly 
better than the other homology modellers (who had access to the same data) when 
judge by independent analysis. There were things wrong with their structures, 
sure, but in almost every case they were less wrong than the other modellers 
(many of whom have been working on this problem for decades).

It _will_ be more impressive once the methods they used (or equivalents) are 
implemented by other groups and are available to the “public” (I haven’t found 
an AlphaFold webserver to submit a sequence to, whereas the other groups in the 
field do make their methods readily available), but it’s still a step-change in 
protein structure prediction - it shows it can be done pretty well.

Michel is right, of course; you can’t have homology modelling without 
homologous models, which are drawn from the PDB - but the other modellers had 
the same access to the PDB (just as we all do…).

Just my two ha’porth.

Harry

> On 8 Dec 2020, at 11:33, Goldman, Adrian 
> mailto:adrian.gold...@helsinki.fi>> wrote:
>
> My impression is that they haven’t published the code, and it is science by 
> press-release.  If one of us tried it, we would - rightly - get hounded out 
> of time.
>
> Adrian
>
>
>
>> On 4 Dec 2020, at 15:57, Michel Fodje 
>> mailto:michel.fo...@lightsource.ca>> wrote:
>>
>> I think the results from AlphaFold2, although exciting and a breakthrough 
>> are being exaggerated just a bit.  We know that all the information required 
>> for the 3D structure is in the sequence. The protein folding problem is 
>> simply how to go from a sequence to the 3D structure. This is not a complex 
>> problem in the sense that cells solve it deterministically.  Thus the 
>> problem is due to lack of understanding and not due to complexity.  
>> AlphaFold and all the others trying to solve this problem are “cheating” in 
>> that they are not just using the sequence, they are using other sequences 
>> like it (multiple-sequence alignments), and they are using all the 
>> structural information contained in the PDB.  All of this information is not 
>> used by the cells.   In short, unless

Re: [ccp4bb] pdb-l: Re: [ccp4bb] Coming July 29: Improved Carbohydrate Data at the PDB -- N-glycans are now separate chains if more than one residue

2020-12-04 Thread Tristan Croll

No - they're changing the auth_asym_id. See 
https://www.wwpdb.org/documentation/carbohydrate-remediation:

Oligosaccharide molecules are classified as a new entity type, branched, 
assigned a unique chain ID (_atom_site.auth_asym_id) and a new mmCIF category 
introduced to define the type of branching (_pdbx_entity_branch.type) .
wwPDB:<https://www.wwpdb.org/documentation/carbohydrate-remediation>
wwPDB: Worldwide Protein Data Bank. Carbohydrate Remediation. As the PDB 
archive grows, and the related science and techniques evolve, the 3D structures 
represented in the Core Archive require ongoing improvement ("remediation") to 
ensure consistency, accuracy, and overall quality.
www.wwpdb.org


From: Greg Couch 
Sent: 04 December 2020 18:51
To: Luca Jovine ; Mailing List CCP4 ; 
Mailing List PDB, 
Cc: Tristan Croll 
Subject: pdb-l: Re: [ccp4bb] Coming July 29: Improved Carbohydrate Data at the 
PDB -- N-glycans are now separate chains if more than one residue

mmCIF has two different chain ids.  One the the label_asym_id which is
used for internal consistency with the entities.  The other is the
auth_asym_id which is whatever the author chooses. If the glycans are
separate entities, then the label_asym_id HAS to be different for each
instance of an entity.  But the auth_asym_id could be the same for all
covalently bonded units.

In ChimeraX, we use the label_seq_id for the chain id when constructing
the molecule and we use the auth_asym_id for the chain id in the user
interface.

 -- Greg




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Re: [ccp4bb] Coming July 29: Improved Carbohydrate Data at the PDB -- N-glycans are now separate chains if more than one residue

2020-12-04 Thread Tristan Croll

OK, I understand your point more clearly now - but I'm not sure I fully agree, 
for the simple reason that people aren't computers. You're right that for the 
purposes of software validation tools the chain IDs are essentially arbitrary - 
as long as they're unique, nothing else really matters. But to a human simply 
wanting to explore a model in their favourite visualisation program this makes 
everything just that bit less intuitive - if they want to, say, go to the first 
glycan attached to chain A they have no way of doing so short of tracing 
through from the N-terminus until they find it, unless the program provides a 
tool that already understands the concept of "first glycan attached to chain 
A". So if we go forward with the "chain IDs are entirely arbitrary, therefore 
it doesn't matter what they are" approach, then every existing visualisation 
tool gets a little bit more difficult to use with glycans until their authors 
take the time to write new task-specific code.

In the grand scheme of things it's a minor issue, I suppose - but in my opinion 
it really is important to keep the experience of the end user in mind when 
making decisions like this.

From: Dale Tronrud 
Sent: 04 December 2020 18:16
To: Tristan Croll ; CCP4BB@JISCMAIL.AC.UK 

Subject: Re: [ccp4bb] Coming July 29: Improved Carbohydrate Data at the PDB -- 
N-glycans are now separate chains if more than one residue

Creating meaning in the chain names "A, B, C, Ag1, Ag2, Ag3" is
exactly the problem.  "chain names" ( or "entity identifiers" if I
recall the mmCIF terminology correctly) are simply database "indexes".
The values of indices are meaningless in themselves, they are just
unique values that can be used to unambiguously identify a record. In
principle, you could just assign random ISO characters (I don't think
mmCIF allows unicode) and the mmCIF would be considered identical.

You are trying to force meaning to the characters with an index, and
that puts multiple types of information in a single field.  As Robbie
said already exists, if you want to encode connectivity into the data
base you have to add records that define that connectivity.  That places
the connectivity information explicitly in the data models and allows
standard data base tools to track and validate.

The idioms of the PDB cause problems that lead people to these
mistakes.  The PDB assigns the indices "1", "2", and "3" to residues in
a chain.  A person could be misled into thinking that "2" comes between
"1" and "3" in the sequence.  This is not necessarily true at all.  To
learn the sequence you have to go to the mmCIF records that define the
connectivity between residues.  It is entirely possible that "3" comes
before "1" because these indexes don't contain any information, other
than being unique within the chain.

Dale Tronrud

On 12/4/2020 9:46 AM, Tristan Croll wrote:
> This suggestion violates a basic principle of data base theory.  A
> single data item cannot encode two pieces of information.
>
> I'm sorry if I was unclear, but I don't believe I was suggesting
> anything of the sort. Hopefully this example should make it more clear -
> I'm just suggesting a slight variation on the existing system, no more:
>
> If we start with model containing 3 protein chains A-C, with chain A
> containing amino acid residues 1-200, and 3 N-linked glycans with
> residues numbered, say, 1000-1005, 1020-1026 and 1040-1043 (a fairly
> common approach I've seen taken to the problem in the past, and one I've
> taken myself), then if I understand correctly after remediation we'll
> have a model with protein chains A-C and glycan chains D-F. The problem
> is, unless and until all the available visualisation software updates to
> automatically associate chains D-F to chain A based on linkage, the user
> just has to remember that chains D-F are actually the chain A glycans.
> This is a simple case, but things quickly become far more messy when you
> have multiple glycosylated species each with multiple glycans per chain.
> If, instead, the new chain assignments were something like "A, B, C,
> Ag1, Ag2, Ag3", then we have something that is far more immediately
> accessible to the user.
>
> 
> *From:* Dale Tronrud 
> *Sent:* 04 December 2020 17:01
> *To:* Tristan Croll ; CCP4BB@JISCMAIL.AC.UK
> 
> *Subject:* Re: [ccp4bb] Coming July 29: Improved Carbohydrate Data at
> the PDB -- N-glycans are now separate chains if more than one residue
>
>  This suggestion violates a basic principle of data base theory.  A
> single data item cannot encode two pieces of information.  The whole
> structure of CIF

Re: [ccp4bb] Coming July 29: Improved Carbohydrate Data at the PDB -- N-glycans are now separate chains if more than one residue

2020-12-04 Thread Tristan Croll

This suggestion violates a basic principle of data base theory.  A
single data item cannot encode two pieces of information.

I'm sorry if I was unclear, but I don't believe I was suggesting anything of 
the sort. Hopefully this example should make it more clear - I'm just 
suggesting a slight variation on the existing system, no more:

If we start with model containing 3 protein chains A-C, with chain A containing 
amino acid residues 1-200, and 3 N-linked glycans with residues numbered, say, 
1000-1005, 1020-1026 and 1040-1043 (a fairly common approach I've seen taken to 
the problem in the past, and one I've taken myself), then if I understand 
correctly after remediation we'll have a model with protein chains A-C and 
glycan chains D-F. The problem is, unless and until all the available 
visualisation software updates to automatically associate chains D-F to chain A 
based on linkage, the user just has to remember that chains D-F are actually 
the chain A glycans. This is a simple case, but things quickly become far more 
messy when you have multiple glycosylated species each with multiple glycans 
per chain. If, instead, the new chain assignments were something like "A, B, C, 
Ag1, Ag2, Ag3", then we have something that is far more immediately accessible 
to the user.

From: Dale Tronrud 
Sent: 04 December 2020 17:01
To: Tristan Croll ; CCP4BB@JISCMAIL.AC.UK 

Subject: Re: [ccp4bb] Coming July 29: Improved Carbohydrate Data at the PDB -- 
N-glycans are now separate chains if more than one residue

This suggestion violates a basic principle of data base theory.  A
single data item cannot encode two pieces of information.  The whole
structure of CIF falls apart if this is done.

Does the new PDB convention contain a CIF record of the link that
bridges between the protein chain and the, now separated, glycan chain?
  If not, I think this is the principle failing of their new scheme.

Dale Tronrud

On 12/4/2020 12:06 AM, Tristan Croll wrote:
> To go one step further: in large, heavily glycosylated multi-chain complexes 
> the assignment of a random new chain ID to each glycan will lead to headaches 
> for people building visualisations using existing viewers, because it loses 
> the easy name-based association of glycan to parent protein chain. A 
> suggestion: why not take full advantage of the mmCIF capability for 
> multi-character chain IDs, and name them by appending characters to the 
> parent chain ID? Using chain A as an example, perhaps the glycans could 
> become Ag1, Ag2, etc.?
>
>> On 4 Dec 2020, at 07:48, Luca Jovine  wrote:
>>
>> CC: pdb-l
>>
>> Dear Zhijie and Robbie,
>>
>> I agree with both of you that the new carbohydrate chain assignment 
>> convention that has been recently adopted by PDB introduces confusion, not 
>> just for PDB-REDO but also - and especially - for end users.
>>
>> Could we kindly ask PDB to improve consistency by either assigning a 
>> separate chain to all covalently attached carbohydrates (regardless of 
>> whether one or more residues have been traced), or reverting to the old 
>> system (where N-/O-glycans inherited the same chain ID of the protein to 
>> which they are attached)? The current hybrid solution hardly seems optimal...
>>
>> Best regards,
>>
>> Luca
>>
>>> On 3 Dec 2020, at 20:17, Robbie Joosten  wrote:
>>>
>>> Dear Zhijie,
>>>
>>> In generally I like the treatment of carbohydrates now as branched 
>>> polymers. I didn't realise there was an exception. It makes sense for 
>>> unlinked carbohydrate ligands, but not for N- or O-glycosylation sites as 
>>> these might change during model building or, in my case, carbohydrate 
>>> rebuilding in PDB-REDO powered by Coot. Thanks for pointing this out.
>>>
>>> Cheers,
>>> Robbie
>>>
>>>> -Original Message-
>>>> From: CCP4 bulletin board  On Behalf Of Zhijie Li
>>>> Sent: Thursday, December 3, 2020 19:52
>>>> To: CCP4BB@JISCMAIL.AC.UK
>>>> Subject: Re: [ccp4bb] Coming July 29: Improved Carbohydrate Data at the
>>>> PDB -- N-glycans are now separate chains if more than one residue
>>>>
>>>> Hi all,
>>>>
>>>> I was confused when I saw mysterious new glycan chains emerging during
>>>> PDB deposition and spent quite some time trying to find out what was
>>>> wrong with my coordinates.  Then it occurred to me that a lot of recent
>>>> structures also had tens of N-glycan chains.  Finally I realized that this
>>>> phenomenon is a consequence of this PDB policy announced here in July.
>>>>
>>>>
>>>&

Re: [ccp4bb] Coming July 29: Improved Carbohydrate Data at the PDB -- N-glycans are now separate chains if more than one residue

2020-12-04 Thread Tristan Croll

To go one step further: in large, heavily glycosylated multi-chain complexes 
the assignment of a random new chain ID to each glycan will lead to headaches 
for people building visualisations using existing viewers, because it loses the 
easy name-based association of glycan to parent protein chain. A suggestion: 
why not take full advantage of the mmCIF capability for multi-character chain 
IDs, and name them by appending characters to the parent chain ID? Using chain 
A as an example, perhaps the glycans could become Ag1, Ag2, etc.?

> On 4 Dec 2020, at 07:48, Luca Jovine  wrote:
> 
> CC: pdb-l
> 
> Dear Zhijie and Robbie,
> 
> I agree with both of you that the new carbohydrate chain assignment 
> convention that has been recently adopted by PDB introduces confusion, not 
> just for PDB-REDO but also - and especially - for end users.
> 
> Could we kindly ask PDB to improve consistency by either assigning a separate 
> chain to all covalently attached carbohydrates (regardless of whether one or 
> more residues have been traced), or reverting to the old system (where 
> N-/O-glycans inherited the same chain ID of the protein to which they are 
> attached)? The current hybrid solution hardly seems optimal...
> 
> Best regards,
> 
> Luca
> 
>> On 3 Dec 2020, at 20:17, Robbie Joosten  wrote:
>> 
>> Dear Zhijie,
>> 
>> In generally I like the treatment of carbohydrates now as branched polymers. 
>> I didn't realise there was an exception. It makes sense for unlinked 
>> carbohydrate ligands, but not for N- or O-glycosylation sites as these might 
>> change during model building or, in my case, carbohydrate rebuilding in 
>> PDB-REDO powered by Coot. Thanks for pointing this out.
>> 
>> Cheers,
>> Robbie
>> 
>>> -Original Message-
>>> From: CCP4 bulletin board  On Behalf Of Zhijie Li
>>> Sent: Thursday, December 3, 2020 19:52
>>> To: CCP4BB@JISCMAIL.AC.UK
>>> Subject: Re: [ccp4bb] Coming July 29: Improved Carbohydrate Data at the
>>> PDB -- N-glycans are now separate chains if more than one residue
>>> 
>>> Hi all,
>>> 
>>> I was confused when I saw mysterious new glycan chains emerging during
>>> PDB deposition and spent quite some time trying to find out what was
>>> wrong with my coordinates.  Then it occurred to me that a lot of recent
>>> structures also had tens of N-glycan chains.  Finally I realized that this
>>> phenomenon is a consequence of this PDB policy announced here in July.
>>> 
>>> 
>>> For future depositors who might also get puzzled, let's put it in a short
>>> sentence:  O- and N-glycans are now separate chains if it they contain more
>>> than one residue; single residues remain with the protein chain.
>>> 
>>> 
>>> https://eur01.safelinks.protection.outlook.com/?url=https%3A%2F%2Fwww.wwpdb.org%2Fdocumentation%2Fcarbohydrate-remediationdata=04%7C01%7Cluca.jovine%40KI.SE%7C1d790a0717ce4217c7a308d897c01b47%7Cbff7eef1cf4b4f32be3da1dda043c05d%7C0%7C1%7C637426199684263065%7CUnknown%7CTWFpbGZsb3d8eyJWIjoiMC4wLjAwMDAiLCJQIjoiV2luMzIiLCJBTiI6Ik1haWwiLCJXVCI6Mn0%3D%7C1000sdata=mBrkCJECFpZyCih4kOCcCvLT1GzQHxD5GD7bZDI9s1s%3Dreserved=0
>>> 
>>> "Oligosaccharide molecules are classified as a new entity type, branched,
>>> assigned a unique chain ID (_atom_site.auth_asym_id) and a new mmCIF
>>> category introduced to define the type of branching
>>> (_pdbx_entity_branch.type) . "
>>> 
>>> 
>>> 
>>> 
>>> 
>>> I found the differential treatment of single-residue glycans and 
>>> multi-residue
>>> glycans not only bit lack of aesthetics but also misleading.  When a 
>>> structure
>>> contains both NAG-NAG... and single NAG on N-glycosylation sites, it might
>>> be because of lack of density for building more residues, or because that
>>> some of the glycosylation sites are now indeed single NAGs (endoH etc.)
>>> while some others are not cleaved due to accessibility issues.Leaving 
>>> NAGs
>>> on the protein chain while assigning NAG-NAG... to a new chain, feels like
>>> suggesting something about their true oligomeric state.
>>> 
>>> 
>>> For example, for cryoEM structures, when one only builds a single NAG at a
>>> site does not necessarily mean that the protein was treated by endoH. In
>>> fact all sites are extended to at least tri-Man in most cases. Then why
>>> keeping some sites associated with the protein chain while others kicked
>>> out?
>>> 
>>> Zhijie
>>> 
>>> 
>>> 
>>> 
>>> 
>>> From: CCP4 bulletin board  on behalf of John
>>> Berrisford 
>>> Sent: Thursday, July 9, 2020 4:39 AM
>>> To: CCP4BB@JISCMAIL.AC.UK 
>>> Subject: [ccp4bb] Coming July 29: Improved Carbohydrate Data at the PDB
>>> 
>>> 
>>> Dear CCP4BB
>>> 
>>> PDB data will shortly incorporate a new data representation for
>>> carbohydrates in PDB entries and reference data that improves the
>>> Findability and Interoperability of these molecules in macromolecular
>>> structures. In order to remediate and improve the representation of
>>> carbohydrates across the archive, the wwPDB has:
>>>

Re: [ccp4bb] AW: [ccp4bb] over-fitting? over-refinement?

2020-10-20 Thread Tristan Croll

I'd like to append a very important caveat to this discussion: most of the talk 
on Rfree as protection against overfitting is perfectly correct, if your 
dataset is high enough resolution. Remember that Rfree only provides protection 
against one form of overfitting: that is, fitting of atoms into random noise. 
What it doesn't protect well against is fitting the wrong atoms into real 
density. Remember, all your x-ray data ultimately says is "there are electrons 
here" - your R-factors don't care where those electrons come from, as long as 
they're present in about the right numbers (with some fudge-room for B-factors 
and occupancies). If you browse through the back catalogue of >3A models, 
you'll find some with horrendous geometry statistics but remarkably good 
R-factors (both work and free) - ultimately, I think, because the model atoms 
have been "overstuffed" into density that is real according to both the working 
and free data. In quite a few such cases I find that even after extensive 
reworking I'm unable to beat the original R-free, despite every other metric 
improving markedly.

Best regards,

Tristan

From: CCP4 bulletin board  on behalf of Barone, Matthias 

Sent: 20 October 2020 12:59
To: CCP4BB@JISCMAIL.AC.UK 
Subject: Re: [ccp4bb] AW: [ccp4bb] over-fitting? over-refinement?


Eleanor rises a very important practical point here..."sidechains at the 
solvent interface have multiple conformations, and that as a result the water 
networks should also have partial occupancies". I was fighting with such a 
model for half a year and also tested XSHEL (there was a thread in here for 
that..). Coupling partial occupancies of sidchains with waters and other 
sidchains is a horrendously time-consuming task...and in the end, as Eleanor 
said, "correcting these details does not change the Rfactors at all". You just 
get fed up with that puzzle and stop right there.

best, matthias


Dr. Matthias Barone

AG Kuehne, Rational Drug Design

Leibniz-Forschungsinstitut für Molekulare Pharmakologie (FMP)
Robert-Rössle-Strasse 10
13125 Berlin

Germany
Phone: +49 (0)30 94793-284


From: CCP4 bulletin board  on behalf of Eleanor Dodson 
<176a9d5ebad7-dmarc-requ...@jiscmail.ac.uk>
Sent: Tuesday, October 20, 2020 12:40:19 PM
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] AW: [ccp4bb] over-fitting? over-refinement?

It is always hard to know when to stop tweaking a model.. We know from high 
resolution studies that many sidechains at the solvent interface have multiple 
conformations, and that as a result the water networks should also have partial 
occupancies. But usually correcting these details does not change the Rfactors 
at all - nor contribute much to the biological relevance of your structure!
So often the point to stop is when you get fed up, Phil Evans said years ago - 
I spend 95% of my time on 5% of the structure, most of which is unimportant..
In practice I let the difference maps decide when to stop - 10 Sigma peak - 
think why - lots of 5 Sigma positive and negative ones not so important
Eleanor

On Tue, 20 Oct 2020 at 11:27, Schreuder, Herman /DE 
mailto:herman.schreu...@sanofi.com>> wrote:

A practice that was very popular before the Rfree came around was to fit a 
water molecule in every noise peak. One would get spectacular low Rfactors this 
way, but I cannot imagine that anyone would believe that this would be fitting 
and not over-fitting.



Best,

Herman



Von: CCP4 bulletin board mailto:CCP4BB@JISCMAIL.AC.UK>> 
Im Auftrag von Sam Tang
Gesendet: Dienstag, 20. Oktober 2020 05:27
An: CCP4BB@JISCMAIL.AC.UK
Betreff: [ccp4bb] over-fitting? over-refinement?



Hi, the question may be a bit weird, but how do you define 'over-fitting' in 
the context of structure refinement? From users' perspective the practical 
aspect is to 'fit' the model into the density. So there comes this question 
from our juniors: fit is fit, how is a model over-fit?



BRS



Sam





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To

Re: [ccp4bb] real real-space-refinement

2020-08-01 Thread Tristan Croll

The isolde suggestion already made is an excellent one. The hardest part of
that is getting the right version of chimeraX working. But, once you've done
that its pretty straightforward.

Hopefully not for too much longer. All going well, ISOLDE 1.0 (to work
in ChimeraX 1.0) should be out this week.

On 2020-07-31 20:33, James Holton wrote:

Not in CCP4, no. And, technically, not in Phenix either. The real-space refinement in Phenix simply picks peaks in the density and then pulls nearby atoms toward them. Like a black hole gobbling up nearby planets. It took me a while to realize that! If you manage to turn off geometry restraints (as I eventually did) all the atoms end up on top of each other. Might seem like a horrible idea, but for poor resolution data and reasonably good geometry restraints it has a high radius of convergence and is incredibly fast when compared to "real real-space refinement". Refining against map voxels directly is a very very slow process.

But, if you real really want to do real real-space, then I suppose coot is
doing that? I'm actually not sure.

The isolde suggestion already made is an excellent one. The hardest part of
that is getting the right version of chimeraX working. But, once you've done
that its pretty straightforward.

One program that has not been mentioned, but does "real real" space refinement is:
"rsref"
https://chapman.missouri.edu/wp-content/uploads/sites/2/software/rsref/html/rsref_doc.html

It is not too hard to install and use. I can't say I've gotten results
appreciably different from reciprocal-space refinement, and that led me to ask
myself why exactly I thought it would be different. The Fourier transform is
symmetric after all. But I do expect that if you have unmodeled regions, such
as big, spiky metals, or large tracts of disordered, ropy stuff, then
localizing the refinement could be beneficial.

Now, of course, you can also do localized refinement in reciprocal space by just smoothing out parts of the map that are "uninteresting". The vast area of noise around the protein in a cryoEM map, for example, is perhaps a candidate for noise suppression. The only trick is how to suppress noise without creating systematic error. For example, if your model does not have "bulk solvent" then this area will be modeled as vacuum, but if you simply set the map voxel values to 0.00, you will have effectively created more bulk solvent, not eliminated it. This is because 0.00 is usually the average voxel value, not the "vacuum level". Then there is the "edge" between the modified and unmodified areas. Unless you smooth it in some way this edge will be very sharp and therefore have significant Fourier coefficients at a wide range of resolutions. So, if you are not careful, your "noise suppression" can create a lot more error than it eliminates.

As for what to do? The scale factor given to the "bulk solvent" model is perhaps the
best value to use to replace the "bulk" solvent region. The bulk solvent mask itself,
ranging from 0 to 1, might also be a reasonable weighting function for combining your original map
with a single-valued map. That is, don't change the protein, but flatten the solvent. You can get
this map out of refmac using the MSKOUT feature. You then smooth it in reciprocal space by applying
refmac's best-fit solvent B factor using sfall and fft, then finally scale it with mapmask. I
should admit, however, that I have not tried this in a while. Let me know if it works!

HTH

-James Holton
MAD Scientist

On 7/29/2020 8:20 AM, Schreuder, Herman /DE wrote:

Dear BB,

I would like to do a real real-space-refinement of a protein against a cryo-EM map; not the mtz-based Refmac approach. A quick internet search produced a lot of Phenix hits, but little ccp4 hits. Does somebody know how to do this using ccp4 programs, or has someone a Coot script to do this?

Thank you for your help!

Herman

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Re: [ccp4bb] Commercial source of His-tagged protein

2020-07-02 Thread Tristan Croll


Thermo Fisher sells His-tagged GFP quite cheaply.
https://www.thermofisher.com/order/catalog/product/A42611#/A42611 


Tristan

On 2020-07-02 10:31, Mark J van Raaij wrote:

Dear All, 

Wondering if anyone knows of an economical commercial source of a soluble His-tagged protein. In principle any His-tagged protein would suffice, because it's for testing coupling to a surface via the His-tag. Asking for an international collaborator, who don't have access/expertise of a molecular biology lab (except us :-).  
We can and will make the His-tagged proteins of interest for them, but if there is an economical and reliable commercial source, it might be worth using that for initial tests. 

Best wishes, 


Mark

Mark J van Raaij
Dpto de Estructura de Macromoleculas
Centro Nacional de Biotecnologia - CSIC
calle Darwin 3
E-28049 Madrid, Spain
tel. (+34) 91 585 4616 
Section Editor Acta Crystallographica F

https://journals.iucr.org/f/

-

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Re: [ccp4bb] disinfecting keyboards

2020-05-06 Thread Tristan Croll

You got off lucky. An old friend of mine learned this lesson when on a 
particularly sunny day he spent an hour out on a New Zealand glacier in 
shorts with no underwear...


On 2020-05-06 16:17, James Holton wrote:

I feel I should correct you on one thing Tim: UV _can_ go around
corners because it scatters.  I learned this the hard way as a younger
man after a fine day of skiing.  I had put sunscreen everywhere except
the bottom of my nose.

You are right, however, that the intensity after scattering is quite a
bit less than tha main illumination.  This is true for all kinds of
light.

-James Holton
MAD Scientist

On 5/5/2020 11:59 PM, Tim Gruene wrote:

Hi James,

for us, the suggestions of cling film / plastic wrap or just swapping
keyboards and mice per person is the simplest - thanks to everyone for
the many suggestions. Especially the latter, since only two people 
will

operate the instruments.

UV light does not go around corners. It might be useful for fume 
hoods,

but for most places, door handles and other curved surfaces are
probably much more the infecting parts, while they escape the UV 
light.


And vira are transported in water droplets, which are larger than 1um.

Best,
Tim

  On Tue, 5 May 2020 17:19:56 -0700
James Holton  wrote:


All joking aside, there has been a furor of attention on UV-based
disinfection of late.  Some of it is not entirely crazy.  I.E.
Columbia University’s Center for Radiological Research has put
forward the idea of illuminating occupied public areas with
ultra-narrow-band UV-C (222 nm).
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5552051/

Mind you, UV-C normally covers 100 - 280 nm, and the PPE requirements
for that (at LBNL at least) are extensive: polycarbonate safety
glasses and face shield with a mark U6 (UV protection), long-sleeved
clothing, and gloves.  Basically: do not expose skin!

The idea behind using monochromatic 222 nm radiation is that it is at
the edge of a very steep increase in the absorption of water,
protein, and other biologicals.  Penetration depths are hard to
estimate because of the steep slope, but they are on the order of 1
micron.  So, smaller than a typical mamalian cell, but bigger than a
bacterium or virus.  The paper above did not have any human subjects,
nor did it discuss how to deal with all the ozone, but the results
are intruiging. Needs further study.

Personally, I think this would probably fog your corneas and perhaps
burn the thin skin on lips and other exposed mucosa. Hair I'd expect
to embrittle and fall apart eventually. Yes, hair is 40 microns thick
and the penetration depth is 1 micron, but photon's don't "stop" at
the penetration depth.  36% of them go deeper. Plastic in keyboards
too would probably bleach and flake with prolonged exposure.  Ever
seen a keyboard left out in the sun for a few weeks?  I'd worry a bit
about this micro-damage creating crevices where bugs could hide.

I encourage you to bring this up with your Health and Safety people,
but make sure they are sitting down first.

-James Holton
MAD Scientist

On 4/29/2020 12:41 PM, Andrea Thorn wrote:

Hi Tim!


100% alcohol is less effective than 80%, and in order to completely
be sure, the keyboard needs not only to be wiped. One can buy
keyboards that can be disinfected because they are waterproof, such
as the Cherry JK-1068DE-2 for about 50 €.


We clean the keyboards in our lab occasionally anyway, and have
used 70% alcohol on them without problem. Disinfectant wipes, a
detergent cleaner (such as Viss Glass & Flächen) and cotton swabs
also offer some help. We wipe our mobile phones with a disinfectant
wipe after washing our hands when arriving home/at work.

I would also be really interested in what could be done with a UV
light, if someone knows?

If the computer is used by one person during the shift, individual
keyboards for each person could be a solution. If people sit down,
the desk surface, which may be touched, should likely also be wiped
at the beginning and end of the shift I would say.

Stay save and best wishes,



Andrea.



Am 29/04/2020 um 21:04 schrieb Diana Tomchick:

100% ethanol or isopropanol work really well on the microscopes,
I soak a Kimwipe and then clean the eyepieces and the knobs for
changing magnification and focus, as well as the door handles,
bench tops, etc.


Diana


**
Diana R. Tomchick
Professor
Departments of Biophysics and Biochemistry
UT Southwestern Medical Center
5323 Harry Hines Blvd.
Rm. ND10.214A
Dallas, TX 75390-8816
diana.tomch...@utsouthwestern.edu
(214) 645-6383 (phone)
(214) 645-6353 (fax)

*From:* CCP4 bulletin board  on behalf of
Diana Tomchick 
*Sent:* Wednesday, April 29, 2020 2:00 PM
*To:* CCP4BB@JISCMAIL.AC.UK
*Subject:* Re: [ccp4bb] disinfecting keyboards

EXTERNAL MAIL

You could try doing what my technician does with her keyboard;
she wraps it in a clear, thin food wrap

Re: [ccp4bb] disinfecting keyboards

2020-04-29 Thread Tristan Croll

Posted semi-seriously (they look like they’d be awful to use, but maybe I’m 
being pessimistic): https://wiki.ezvid.com/best-virtual-keyboards. With a 
projection keyboard, all you have to worry about cleaning is the benchtop.
 

 

> On 29 Apr 2020, at 21:38, Crissy Lynette Tarver  wrote:
> 
> https://pdb101.rcsb.org/learn/videos/fighting-coronavirus-with-soap
> 
> 
> Crissy L Tarver
> Postdoctoral Researcher
> Department of Structural Biology
> Stanford University School of Medicine
> From: CCP4 bulletin board  on behalf of Tim Gruene 
> 
> Sent: Wednesday, April 29, 2020 11:53:32 AM
> To: CCP4BB@JISCMAIL.AC.UK 
> Subject: [ccp4bb] disinfecting keyboards
>  
> Dear all,
> 
> can you make suggestions for how to disinfect computer keyboards, and
> instrument panels?
> 
> Our facility is going to reboot next week, with shifts so that people
> don't meet. The main interface will be the computer keyboards, as well
> as the door of our X-ray diffractometer and the mounting of the
> crystals.
> 
> The keyboard labels may not like alcohols (and the efficiency of
> injecting disinfecting through the USB cable is also under discussion,
> so I heard).
> 
> One way would be to use individual keyboards, and wearing gloves for
> replugging, and to use gloves for mounting crystals.
> 
> But maybe there are other ways that won't require gloves?
> 
> Best regards,
> Tim
> 
> -- 
> --
> Tim Gruene
> Head of the Centre for X-ray Structure Analysis
> Faculty of Chemistry
> University of Vienna
> 
> Phone: +43-1-4277-70202
> 
> GPG Key ID = A46BEE1A
> 
> 
> 
> To unsubscribe from the CCP4BB list, click the following link:
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Re: [ccp4bb] disinfecting keyboards

2020-04-29 Thread Tristan Croll

I once knew a particularly germaphobic IT manager who used to absolutely 
*everything* via ssh or remote desktop from his own laptop, even when he 
was physically in front of the user's computer. If feasible, that 
approach would actually seem quite smart in the current environment.


On 2020-04-29 19:53, Tim Gruene wrote:

Dear all,

can you make suggestions for how to disinfect computer keyboards, and
instrument panels?

Our facility is going to reboot next week, with shifts so that people
don't meet. The main interface will be the computer keyboards, as well
as the door of our X-ray diffractometer and the mounting of the
crystals.

The keyboard labels may not like alcohols (and the efficiency of
injecting disinfecting through the USB cable is also under discussion,
so I heard).

One way would be to use individual keyboards, and wearing gloves for
replugging, and to use gloves for mounting crystals.

But maybe there are other ways that won't require gloves?

Best regards,
Tim

--
--
Tim Gruene
Head of the Centre for X-ray Structure Analysis
Faculty of Chemistry
University of Vienna

Phone: +43-1-4277-70202

GPG Key ID = A46BEE1A



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[ccp4bb] ISOLDE 1.0b5 release

2020-04-06 Thread Tristan Croll


Hi all,

For those still lucky enough to be working on molecular model building 
during this crisis, I've just pushed ISOLDE 1.0b5 out to the ChimeraX 
ToolShed (to work with the ChimeraX 0.93 release version - download from 
http://preview.cgl.ucsf.edu/chimerax/download.html#release, then install 
ISOLDE from within ChimeraX via Tools/More Tools...). While development 
has been a little light since version 1.0b4 due to travel in February 
and then, well... everything since, there are a few new features and 
bug-fixes worth downloading for:


- Perhaps the biggest improvement isn't actually mine - thanks to some 
stellar work by Tom Goddard at UCSF, ChimeraX's ribbon drawing is now 
ten times faster than before. Particularly for those working with very 
large models, this drastically reduces the lag time upon finishing an 
interactive simulation and returning to the default display mode.


- I've done some optimisations to the methods for choosing the initial 
map contour levels and the MDFF coupling constant (how strongly the map 
"pulls" on the model during simulations). They're now about 20 times 
faster, removing most of the delay on initial preparation of a model for 
ISOLDE.


- If you're lucky enough to be working on a multi-GPU machine, you can 
now explicitly specify the GPU to use for simulations with the command 
"isolde set gpuDeviceIndex {a number}" (you may need to experiment to 
work out which GPU is which)


- You can adjust the number of simulation steps between GUI updates with 
"isolde set timeStepsPerGuiUpdate". This is particularly useful for 
slower GPUs - your simulation won't actually run faster, but your 
experience of it will be much smoother. The default value of 50 works 
well for high-end GPUs; for (for example) light-weight MacBook GPUs a 
value of 10 may be more suitable. Performance curves for a few of the 
machines available to me can be found at 
https://isolde.cimr.cam.ac.uk/performance-benchmarks/.


- The command "clipper symmetry #{model number}" will draw a unit cell 
with all rotational symmetry and screw axes (mirror symmetry is not yet 
handled). The depiction is still a work in progress, but the 
fundamentals are there.


- While not new to 1.0b5, a reminder that since 1.0b4 ISOLDE is fully 
compatible with ChimeraX's session save/restore. The command "save 
{filename}.cxs" will save your current session (including your model(s), 
all custom restraints, maps, weightings and visualisation state) to a 
file which can be reopened on the same or another machine to pick up 
exactly where you left off.



Known bug: (seemingly only) on Linux machines, certain links to the help 
files (e.g. by typing "usage isolde" on the command line and then 
clicking the "isolde adjust distances"link in the log) sometimes cause 
segmentation faults if the help browser is not already open, crashing 
ChimeraX. This appears to be a bug in the Qt 5.12 web browser 
implementation, but so far has proven near-impossible to isolate. To 
avoid it, simply make sure the help browser is open (Help/User Guide) 
before clicking any links in the log.



I very much hope you find it useful.

Stay healthy!

Tristan



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Re: [ccp4bb] CCP4BB vs COVID19

2020-03-22 Thread Tristan Croll


For what it’s worth, I’ve spent the last few days going over most of the 
existing COVID-19 related structures and rebuilding/re-refining wherever I 
considered necessary. The resulting models along with some basic explanatory 
notes are at 
https://drive.google.com/drive/folders/1S5qJtCnK00NrcbwwBNgImUMewhiBkyPa?usp=sharing.
 Some commentary on things I found can also be found in my Twitter thread at 
https://twitter.com/crolltristan/status/1241317484235554823?s=21. Normally I 
shy away from meddling with other people’s new models without invitation - but 
if ever there was a time to make an exception, this is it. Please feel free to 
use these for whatever purpose you like, and to improve on them further if you 
can. Tomorrow I’ll start personally contacting the individual authors to see if 
they’re willing to update their PDB entries.

Wishing the best of good health to all!

Tristan
 

 

> On 20 Mar 2020, at 22:59, James Holton  wrote:
> 
> You might think that as a structural biologist you won't be able to do much 
> about COVID-19 anytime soon, but that is not true.  Yes, real-world 
> therapeutics and vaccines take time, but we have already seen just how fast 
> we can get started.  There are 21 PDBs already and some even have bound 
> ligands.  Good job Frank et al. BTW!  And my personal thanks to all of you 
> out there who are already hard at work on this.
> 
> I believe this forum is an ideal place to share information and ideas on the 
> structural biology of SARS-CoV-2 as we move forward. It's a big virus, but 
> there are not that many proteins in it.  If all of us independently do the 
> same bioinformatics and literature searches and end up trying exactly the 
> same thing in every lab all over the world, then that would be more than 
> unfortunate.  To that end, I am personally interested on ORF8 for reasons I 
> will go into below.  Has anyone tried to solve it yet?  What happened?  
> Didn't express? Bad diffraction?  What?  Do tell.
> 
> Some of us, as you may have heard, are stuck at home, our beamlines and labs 
> dark while we shelter-in-place.  That doesn't mean our hands are tied.  We 
> are still allowed to think. The fraction of the human race that has a 
> snowball's chance in Hades of figuring out this bug is very very small.  
> Structure may be your main skill set, but you are still a biologist.  Do you 
> know how to run a PCR machine?  Do you know how to pipette?  You might think 
> that anybody can do it, but that is really not the case. Ever trained a new 
> student on sterile technique?  How many days did that take?  Now remember 
> that your student was no dummy and already studying biology.  Everyone 
> reading this will make an excellent volenteer at the very least.  I'm not 
> saying this to belittle the average human, only to say that we scientists, 
> moving in the circles we do, often forget that we have uncommon capabilities.
> 
> For example, I also believe we can be useful in assay development. The void 
> left by the dearth and delay of test results has been filled with fear, and 
> that is a big problem.  The tests, as defined, are straightforward, but also 
> extremely regimented like any good laboratory protocol should be.  The US 
> CDC's instructions for academic labs are here:
> https://www.cdc.gov/coronavirus/2019-nCoV/lab/index.html
> My question is: how can this test be made faster, using more commonplace 
> supplies, in high-throughput mode and still valid?  Not just for clinical but 
> for academic use?  I think more than a few people on this list could be 
> regarded as experts in making a complex biochemical task faster, more 
> efficient, high-throughput and nonetheless valid.  Yes, there are other 
> people who do virus testing for a living, but right now they are all rather 
> busy.  Maybe if we put our minds to it we can help?
> 
> As for why ORF8.  I am basing my interest on the bioinformatics done in this 
> article: https://dx.doi.org/10.1093/nsr/nwaa036.  Search for "T8517C" and you 
> will find what I'm talking about.  The authors found two "types" of 
> SARS-CoV-2.  They call them "S" and "L" because the only conserved amino acid 
> change involved is S84L in ORF8.  The "S" type is believed to be the ancestor 
> of "L".  What is interesting is how tightly linked this mutation is to a 
> silent mutation on the other end of the genome: the "L" type has a faster 
> codon for Ser in ORF1.  Such tight coupling (r^2=0.945) means there must be 
> significant selective pressure preventing both of these mutations occurring 
> in the same virus at the same time.  That, I believe, is interesting.  
> Espeically since they are so far apart I expect this selective pressure might 
> work in trans: as in a super-infection. That is, the S and L genome types may 
> interfere with each other.
> 
> The authors fall short of claiming evidence of interference upon 
> super-infection, and indeed they have already been criticised for calling "L" 
> the

Re: [ccp4bb] Generating symmetry mates using python

2020-01-12 Thread Tristan Croll

Just to add to the already-excellent list of replies, this can also be done 
quite straightforwardly with the Clipper plugin in ChimeraX. I’d be happy to 
provide an example script if you want one.

Best regards,

Tristan
 

 

> On 12 Jan 2020, at 18:54, orly avraham  wrote:
> 
> Hi Marcin,
> 
> Thank you very much for your detailed reply!
> You pointed out many special cases that I wouldn't have thought of.
> Indeed an existing solution is highly preferred, and I appreciate you 
> pointing out several available libraries. I came across Gemmi and I will give 
> it another look, and I will examine the additional resources you mentioned as 
> well.
> 
> Best regards,
> Orly Avraham
> 
>> On Sun, Jan 12, 2020 at 5:59 PM Marcin Wojdyr  wrote:
>> Hi Zhijie,
>> 
>> it's good and instructive to implement such things from the ground up,
>> but there are many special cases that one would be discovering while
>> testing this procedure, so if the time is limited it may be better to
>> use an existing solution.
>> 
>> For instance, here one may find out that using the SCALE1 record
>> doesn't give the sufficient accuracy. In the example in your script
>> you have 6 significant digits in the unit cell lengths in CRYST1, but
>> only 4 significant digits in SCALE1. (The accuracy of SCALE1 is
>> problematic in general; sometimes it needs to be manually removed when
>> a program reads it in preference to CRYST1.)
>> 
>> Then one may find out that the 3x3x3 supercell is not sufficient. If
>> the molecule is far from the origin, symmetry operations send it far
>> away. For example, 5M3H annotates (in the mmCIF format) hydrogen bond
>> between 1_555 and 2_11516 - the symmetry mate is shifted 16 unit cells
>> in the z direction. Since you already use fractional coordinates in
>> your script you could tell directly from the center-of-mass
>> coordinates how many unit cells it should be shifted. Say, you have
>> x=3.1, so to shift it near the origin you shift it by 3 unit cells
>> along x.
>> 
>> But even if all the molecules are shifted near the origin, the 3x3x3
>> cell is still not sufficient to find contacts.
>> See 3NWH – a homo-4-mer in P2 (4 x 2 chains per unit cell). Here it in
>> its unit cell, colored by the chain id:
>> https://gemmi.readthedocs.io/en/latest/_images/3nwh.png
>> Or 5XG2 – a monomer in P21. Two copies of the chain are rainbow-colored here:
>> https://gemmi.readthedocs.io/en/latest/_images/5xg2.png
>> These chains span over more than 4 unit cells in one direction. One
>> could use big enough supercell, but it'd be slow. I suppose that even
>> using a 3x3x3 supercell is slow. The alternative is to do the distance
>> calculation in fractional coordinates modulo 1.
>> 
>> Then you needs to consider atoms on special positions. If you apply
>> symmetry operations to an atom on a 4-fold symmetry axis you get 4
>> atoms in the same place. So this needs to be handled. The atom may not
>> be exactly on the axis, because the refinement program may not
>> constrain its position. So the symmetry operations should produce, I
>> think, 4 alternative locations of the same atom. But you could also
>> have an atom near the symmetry axis bonded to its symmetry mate - then
>> the symmetry operations should produce different atoms. So the
>> procedure requires a cut-off distance or a heuristics to distinguish
>> the two cases.
>> 
>> Then, if you'd like to expand non-crystallographic symmetry from the
>> MTRIX records - this is another complication. And so on...
>> 
>> So I'd recommend using one of many available programs for finding
>> contacts or interactions. If none of them is suitable - then try
>> crystallographic libraries.
>> I didn't document yet how to find the contacts using gemmi, but I'll
>> do it in the coming weeks (or months). Cctbx and clipper are other
>> (more mature) libraries worth checking.
>> 
>> Best wishes,
>> Marcin
>> 
>> 
>> On Sat, 11 Jan 2020 at 02:11, Zhijie Li  wrote:
>> >
>> > Hi Orly,
>> >
>> > REMARK 290 should be the easiest way for generating symmetry mates. Other 
>> > routes are just going to give you the same results. As Jonathan already 
>> > pointed out, the symm ops do not garantee that the symm copies are close 
>> > to each other.  The most simple-minded solution to this problem would be 
>> > simply generating 3x3x3 unit cells so that the unit cell in center will be 
>> > complete. An upgrade to this is to compute the center of mass of the 
>> > symmetry copies in each of the 3x3c3 cells and find which one is closest 
>> > to the orignal 1555 copy.  Just for fun, I wrote a little python script 
>> > that does this (attached). In this script for unit cell translation and 
>> > calculating center-center distances, I converted the Cartesian coordinates 
>> > to fractional coords first. Then after the translation,I used the inverse 
>> > of the SCALE1 matrix to get the shifted Cartesian coords. This way I don't 
>> > need to read wikipedia on geometry . But as noted in the script

[ccp4bb] ISOLDE 1.0b4 released

2020-01-03 Thread Tristan Croll

Happy new year to all my colleagues in the structural biology community! 
I'm delighted to announce the release of ISOLDE 1.0b4 to coincide with 
the stable release of ChimeraX 0.91.


What is ISOLDE?

ISOLDE is a plugin to UCSF ChimeraX, designed to ease the task of 
macromolecular model building into low-to-medium resolution maps derived 
from crystallographic or electron cryo-microscopy (cryoEM) experiments. 
Historically, this has been an extremely challenging task, since at low 
resolutions the maps alone are insufficient to precisely place 
individual atoms. ISOLDE aims to reduce this challenge in a number of 
ways:


- Rebuilding is accomplished via GPU-accelerated interactive molecular 
dynamics (using OpenMM and the AMBER molecular dynamics forcefield) to 
make the task feel as close as possible to what it might be like to work 
with a real physical molecule.
- Geometric validation of protein backbone and sidechain conformations 
is performed in real time, allowing you to see problem sites directly on 
the model as you work with it.
- Remodelling can be performed by directly tugging on atoms, or via the 
interactive addition and removal of position, torsion and/or distance 
restraints
- For crystallographic datasets, structure factors are constantly 
recalculated in the background as the model coordinates change - as the 
model improves, you see the map improve.



For more details, see https://isolde.cimr.cam.ac.uk/.

To install it for Linux, MacOS or Windows 10, go to 
http://preview.cgl.ucsf.edu/chimerax/download.html#release and install 
the **0.91** release build for your operating system.


(IMPORTANT NOTES: this build of ISOLDE will **only** work with version 
0.91 of ChimeraX. Additionally, Ubuntu users should use the "Generic 
Linux" ChimeraX build - the Ubuntu builds use compilers that are not 
compatible with ISOLDE).


Start ChimeraX, go to Tools/More Tools... and follow the links to 
install ISOLDE. Equivalently, just type "toolshed install isolde" on the 
ChimeraX command line. You can then start ISOLDE via 
Tools/General/ISOLDE or the command "isolde start". If this is your 
first time using it I highly recommend running either or both of the 
first two tutorials found in the top left of the ISOLDE panel that will 
appear.


A short list of what's new since 1.0b3:

- source code for ISOLDE and its "sister" package ChimeraX-Clipper are 
now on GitHub at https://github.com/tristanic/isolde and 
https://github.com/tristanic/chimerax-clipper respectively.
- integration with ChimeraX session save/restore framework. "save 
session.cxs" will save a snapshot of all models, maps and strucuture 
factors including all current ISOLDE restraints, and "load session.cxs" 
will return you to where you left off.
- added new adaptive torsion restraint implementation (a periodic 
analogue to Geman-McClure distance restraints) to restrain protein 
torsions to their current geometry or to a reference model. For details, 
type "usage isolde restrain torsions" on the ChimeraX command line and 
click the link in the log.
- documentation is now integrated with the ChimeraX documentation 
framework. In particular, "usage {command}" now provides a link to full 
documentation in the ChimeraX log window.

- improved behaviour (speed and stability) of energy minimiser
- significantly improved initial startup and simulation start times
- clearer indication of simulation state (running/paused)
- remote Python interface for communication with other packages
- various improvements and bug-fixes to handling of crystallographic 
data. In particular: better handling of anisotropic datasets, and 
improved stability of bulk solvent and scaling calculations.

- improved performance of live crystallographic map updates.
- support for multiple independent sets of adaptive distance restraints 
(currently only available via the Python API). For example, one set may 
be used to maintain internal geometry, while another represents 
crosslinking/mass spec or evolutionary coupling restraints.
- "Demo" button on ISOLDE GUI retired and replaced with a "Tutorials" 
drop-down menu.
- added tugging support in ChimeraX VR (thanks to Tom Goddard). ChimeraX 
currently supports VR only in Windows.




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Re: [ccp4bb] Changes in quaternary structure

2019-10-09 Thread Tristan Croll

For that matter, the insulin *receptor* is a spectacular example of very 
large conformational changes in a homodimer.


On 2019-10-09 15:27, Eleanor Dodson wrote:

There are many insulin structures with domain shifts.
4ins and 1trz share sequence but have different conformations for one
of the molecules..
Eleanor Dodson

On Wed, 9 Oct 2019 at 14:32, Tanner, John J. 
wrote:


The same enzyme adopting different quaternary structures is a form
of allosteric regulation. Jaffe's group has described this
phenomenon - the “morpheein" model of allosteric regulation.

Reviews:
https://www.ncbi.nlm.nih.gov/pubmed/22182754 [2]
https://www.ncbi.nlm.nih.gov/pubmed/16023348 [3]

John J. Tanner, PhD
Professor of Biochemistry and Chemistry
Associate Chair of Biochemistry
University of Missouri
117 Schweitzer Hall
503 S. College Avenue
Columbia, MO 65211
Phone: 573-884-1280
Email: tanne...@missouri.edu
http://faculty.missouri.edu/~tannerjj/tannergroup/tanner.html [4]


On Oct 9, 2019, at 6:14 AM, Gabriela GARCIA RODRIGUEZ
 wrote:

Dear CCP4BB subscribers,

I am writing a discussion on extensive quaternary changes between
two different homodimers of the same protein and I would like to
ask you for any examples of your own work/that you know of to
include in my analysis. Basically, any proteins known to exist in
two different (or more) homodimerised states (suffering changes in
domain organisation between states would be even more
interesting).
I would greatly appreciate any examples! Thank you for your time
and attention.

Warm regards,

Gabriela



GABRIELA GARCIA RODRIGUEZ

_gabriela.garcia.rodriguez@vub.be_

PhD candidate

Structural Biology Research Centre (SBRC)

Vlaams Instituut voor Biotechnologie (VIB) - Vrije Universiteit
Brussel (VUB)

Pleinlaan 2- 1050 Brussels, Belgium

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Re: [ccp4bb] I am doing phenix refinement now. Is it a problem if the r-work and r-free values are equal?

2019-08-30 Thread Tristan Croll

... although this raises a question: if there are no free reflections, 
then why is a "Rfree" score being reported at all?


On 2019-08-30 16:53, Christian Roth wrote:

No.  As pointed out by Pavel, there are no reflections flagged as
"free reflections" used to calculate Rfree. Therefore you don't get a
real Rfree. No model bias issue etc. If you have a Rfree using ccp4
programs, than you should used that particular original mtz in phenix.
It should detect the flagged "free reflections" automatically.

Cheers
Christian

On Fri, Aug 30, 2019 at 5:24 PM Tung Thanh Dinh 
wrote:


I have been using elipsoidal truncated data all long. And at first i
couldn't find a well homologous model (50% homology is the best i
can find) so I use Mr Bump and Buccaneer to create a model to phase
and autobuild. After a lot of refinement I obtained "good" model
with good statistics. Then I used this "good" model to either phase
and autobuild again. So this may be due to the model bias?

Kindest regards,
Tung Dinh

PhD student
The University of Georgia
Department of Chemistry
-

FROM: CCP4 bulletin board  on behalf of Tung
Thanh Dinh 
SENT: Friday, August 30, 2019 9:36:58 AM
TO: CCP4BB@JISCMAIL.AC.UK 
SUBJECT: [ccp4bb] I am doing phenix refinement now. Is it a problem
if the r-work and r-free values are equal?

After phasing with phenix, i clicked on the phenix.refine ribbon.
Since then for every macrocycle of refinement, r-work and r-free
values are always the same. Is this a problem that I need to fix?

Kindest regards,
Tung Dinh

PhD student
The University of Georgia
Department of Chemistry

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Re: [ccp4bb] [OT] Structure-related pun needed urgently

2019-08-15 Thread Tristan Croll


Everyone keeps telling me I look terrible, but I'm unphased...


On 2019-08-15 12:42, Gerard DVD Kleywegt wrote:

Dear CCP4BB-ers,

Once again I turn to you in my hour of need. I *urgently* need a
side-splittingly funny, and ideally punny, structure-related
sentence/statement/claim/expression to put in a speech bubble attached
to a life-size bobblehead version of yours truly (don't ask)!

I know there are some very funny people on CCP4BB. The best I've been
able to come up with myself so far is: "Protein structures are
beautiful, but I try to keep it platomic" - which is pretty lame, I
know.

Many thanks in advance!

--Gerard

**
   Gerard J. Kleywegt

  http://xray.bmc.uu.se/gerard   mailto:ger...@xray.bmc.uu.se
**
   The opinions in this message are fictional.  Any similarity
   to actual opinions, living or dead, is purely coincidental.
**
   Little known gastromathematical curiosity: let "z" be the
   radius and "a" the thickness of a pizza. Then the volume
of that pizza is equal to pi*z*z*a !
**



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[ccp4bb] ISOLDE dev builds now available

2019-07-30 Thread Tristan Croll


Hi all,

Since the release of ISOLDE 1.0b3 I've begun releasing regular 
(weekly~fortnightly) development builds of ISOLDE. These are built 
against the ChimeraX daily build at the time of release - if you 
download a ChimeraX-daily build and install ISOLDE in the usual way via 
Tools/More Tools... this is what you'll get. In case you're worried, the 
dev and official 1.0b3 builds install into different directories, so 
it's quite straightforward to have both versions running independently 
on the same machine. The dev builds are *not* compatible with the 
ChimeraX 0.9 stable release.


Some important changes since 1.0b3/ChimeraX 0.9:

- the ChimeraX team have fixed some bugs affecting PDB and mmCIF I/O 
(primarily involving the handling of "imperfect" files)
- fixed a bug causing the simulation to crash if the minimiser didn't 
converge in the first round
- added status messages during simulation startup, and clear indications 
of when a simulation is running/paused
- improved simulation startup time, and handling of custom ligands 
(preparing ligand definition files is a work in progress and will 
currently need you to do some work outside ISOLDE - feel free to contact 
me if you have an urgent case)


Best regards,

Tristan



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Re: [ccp4bb] AW: [EXTERNAL] [ccp4bb] Questionable Ligand Density: 6MO0, 6MO1, 6MO2

2019-07-19 Thread Tristan Croll

I couldn't resist doing a little playing with 6mo0 in ISOLDE. The
sequence past Gly1151 is wrong - after jumping a small gap (density
clearly indicates a single missing residue here), it continues on from
1165. Corrected the following sequence to GVVTRS, deleted the ligands,
did some basic energy minimisation and cleaning up of obvious local
problems, reset the B-factors and refined in Phenix.refine. Rwork/Rfree
28/32, MolProbity score 2.14 (vs. 3.6 originally). No sign of anything
like the ligand in the difference density (just a small blob in the
active site). Some other oddities make me suspect a number of point
mutations from the assigned sequence (not register errors - the
surrounds are quite clear). I'm particularly intrigued by residues 48-71
- this stretch appears to link seamlessly head-to-tail with its own
symmetry copy, forming a seemingly-continuous chain snaking through the
crystal. Can't make head nor tail of that. A decidedly odd structure...
but I agree with you that the ligand doesn't seem supported by the
density.

Best regards,

Tristan

On 2019-07-19 14:46, herman.schreu...@sanofi.com wrote:

Hi Rhys,

There is definitively some density present for a ligand, but the
active site region looks completely misfitted, and the ligand density
may also belong to unfitted protein residues. One first needs to get
the protein chain right, and should then look if there would still be
density available to fit the ligand.

Best,

Herman

VON: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] IM AUFTRAG VON
Rhys Grinter
GESENDET: Freitag, 19. Juli 2019 15:22
AN: CCP4BB@JISCMAIL.AC.UK
BETREFF: [EXTERNAL] [ccp4bb] Questionable Ligand Density: 6MO0, 6MO1,
6MO2

EXTERNAL : Real sender is owner-ccp...@jiscmail.ac.uk

Hi All,

I was chatting with a colleague during a recent synchrotron visit and
they'd recently come across some ligand/drug bound structures
associated with a paper recently published in a high impact factor
journal.

They had pulled the associated SFs from the PDB and found that the
electron density associated with these ligands didn't match that
reported in the paper and certainly wasn't sufficient to model the
alleged ligand.

I also pulled the structure factors and after refinement in the
presence/absence of the alleged ligand I also feel that the density
present does not warrant modelling of the ligand.

I was hoping that the community might be able to give me an outside
opinion on these datasets (PDB IDs: 6MO0, 6MO1, 6MO2) and if the
problem associated with the data is verified, provide some advice on
how to proceed.

This isn't the first occasion I've seen ligand bound structures with
questionable density deposited in association with papers in well
respected journals. Despite improvements to validation I feel that
this problem is widespread.

Best Regards,

Rhys

Dr Rhys Grinter

NHMRC Postdoctoral Researcher

Monash University

+61 (0)3 9902 9213

+61 (0)403 896 767

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Re: [ccp4bb] challenges in structural biology

2019-07-17 Thread Tristan Croll


Hi Radu,

Barring some truly spectacular advances, I think that crystallography is 
going to have a major role to play for a long time yet. Looking at 
single-particle cryoEM, for almost every target apart from the few 
ultra-rigid "rocks" the reconstruction will have a wide range of 
resolutions from (near)atomic in the rigid core to fuzzy blobs out on 
the floppy exterior. Yes, modern focused reconstruction techniques are 
improving upon this all the time, but there will be limits - and the 
problem is much more serious when it comes to electron cryo-tomography 
methods where you can't simply collect more particles! I think there's 
some lovely potential complementarity between cryo-EM and 
crystallography here: almost by definition, these peripheral mobile 
domains tend to be things that fold independently of the main body of 
the complex - so why not express them independently and see if they 
crystallise? That way you get the best of both worlds: the initial 
cryo-EM reconstruction allows some informed decision making on what 
construct(s) is/are likely to reliably crystallise, crystallisation of 
those domains gives you the atomic-resolution description you need, and 
modelling these back into the cryo-EM map allows you to study them in a 
more natural context.


To bring things back to the original topic, I guess that's what I'd like 
to see more of: rather than seeing the methods as in fundamental 
competition, where are the *complementarities* between crystallography 
and newer techniques?


Best regards,

Tristan

On 2019-07-17 08:43, r...@mrc-lmb.cam.ac.uk wrote:

Hi Both,

I am not questioning the PDB stats, the issue was whether (crystal) 
structures
are sufficiently relevant to address biological questions and justify 
the
resources. Fragment screening is one example where investment in 
protein
crystallography can still be justified (for now). But it doesn't really 
ask or

answer biological questions... for these, whether we like it or not,
macromolecular crystallography (or NMR, even in cell) cannot be the 
future. In

my opinion :-)

Best wishes,

Radu


Stating the crystallography is dead might be a bit premature, it is 
still king

for depositions.



In 2017 we had a large number of fragment screening experiments 
deposited.








From: CCP4 bulletin board  On Behalf Of Nukri
Sanishvili
Sent: 15 July 2019 23:09
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] challenges in structural biology



I know it is going to hijack the original topic but I could not 
help...




â€œThe reports of death of (macromolecular) crystallography are 
greatly

exaggerated.

If we believed the prognosticators, it has been dead since the 80s 
when some
folks made the claim that the only relevant structures were those 
solved by

NMR.

I think we've done quite well since then...

Best,

Nukri



On Mon, Jul 15, 2019 at 3:45 PM mailto:r...@mrc-lmb.cam.ac.uk> > wrote:

Hi Tassos, Tim,

I wonder why would you or anyone on this list worry whether biological
questions that can be asked and answered with structures are relevant 
to
justify the resources? I think there is abundant evidence that this is 
the
case. Unless your point is that crystallography is now dead for all 
practical
purposes... then yes, I fully agree :-) It would however be wrong to 
erase its

historical contribution to understanding biology.

Best wishes,

Radu



I would wonder more if the biological questions you can *ask* with a
(crystal)
structure are sufficiently relevant to justify the resources.

Sent from my iPhone


On 15 Jul 2019, at 22:08, Tim GrÃ¼ne mailto:tim.gru...@univie.ac.at> > wrote:

Dear James,

10) are the biological questions that you can answer with a 
(crystal)

structure sufficiently relevant to justify the resources?

Best,
Tim



Am 15.07.2019 21:44, schrieb Holton, James M:

Hello folks,
I have the distinct honor of chairing the next Gordon Research
Conference on Diffraction Methods in Structural Biology (July 26-31
2020).  This meeting will focus on the biggest challenges currently
faced by structural biologists, and I mean actual real-world
challenges.  As much as possible, these challenges will take the 
form of
friendly competitions with defined parameters, data, a scoring 
system,
and "winners", to be established along with other unpublished 
results

only at the meeting, as is tradition at GRCs.
But what are the principle challenges in biological structure
determination today?  I of course have my own ideas, but I feel 
like I'm

forgetting something.  Obvious choices are:
1) getting crystals to diffract better
2) building models into low-resolution maps (after failing at #1)
3) telling if a ligand is really there or not
4) the phase problem (dealing with weak signal, twinning and
pseudotranslation)
5) what does "resolution" really mean?
6) why are macromolecular R factors so much higher than 
small-molecule

ones?
7) what is the best way to process serial crystallography data?
8) how should one deal with

[ccp4bb] ISOLDE 1.0b3 release

2019-06-17 Thread Tristan Croll


Dear structural biology community members,

I am happy to announce that ISOLDE 1.0b3 is now live and available for 
installation via the ChimeraX Tool Shed. To get it, just download, 
install and run the ChimeraX 0.9 release version from 
http://preview.cgl.ucsf.edu/chimerax/download.html#release, then go to 
Tools/More tools… and find ISOLDE in the web page that appears.


If you’re unfamiliar with what ISOLDE is about, in brief it’s an 
interactive model-building environment specifically tailored for the 
challenges encountered in fitting low-medium resolution maps. Find out 
more at https://isolde.cimr.cam.ac.uk.


Best regards,

Tristan

What’s new in 1.0b3:

- parallelised structure factor calculations (courtesy of ISOLDE’s 
sister plugin, ChimeraX_Clipper).
- support for a wider range of chemical space (glycosylations courtesy 
of GLYCAM and various other post-translational modifications, and 
~13,000 of the more common ligands in the wwPDB thanks to the work of 
Nigel Moriarty and David Case)
- adaptive distance restraints (for reference model restraints and/or 
large-scale interactive flexible fitting – see 
https://isolde.cimr.cam.ac.uk/features-main/interactive-restraints/).
- ability to fetch model and structure factors (as F/sigF, I/sigI or 
anomalous equivalents) straight from the wwPDB
- interactive tutorials (run the command “isolde tut” on the ChimeraX 
command line)

- various general usability and performance improvements.



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Re: [ccp4bb] MolProbity website down

2019-06-04 Thread Tristan Croll

Seems to be working fine: 
https://downforeveryoneorjustme.com/molprobity.biochem.duke.edu. It's 
not a secure site, though: https://molprobity... times out, whereas 
http://molprobity... goes through. Could that be your issue?


On 2019-06-04 13:19, Andrea Pica wrote:

Hi everyone,

it seems that MolProbity website (molprobity.biochem.duke.edu) is not 
reachable.


Anyone knows what happened?

Cheers,

Andrea


--
Andrea Pica. Ph.D.
Postdoctoral Researcher
High-Throughput Crystallization Lab
EMBL Grenoble Outstation
Postal address: European Molecular Biology Laboratory
71, Avenue des Martyrs
CS 90181 38042 Grenoble Cedex 9, France
Delivery address: European Molecular Biology Laboratory
71, Avenue des Martyrs
38000 Grenoble, France
Phone +33 (0) 47 620 7632



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Re: [ccp4bb] select from a pdb file a sphere of atoms

2019-04-03 Thread Tristan Croll


In ChimeraX, you can promote a selection to whole residues with:

select up

Repeating the command will expand to whole secondary structure elements, 
then whole chains, then whole models. You can reverse this with:


select down


On 2019-04-03 16:13, Gianluca Santoni wrote:

I could be useful to then expand the selection to include the full
residues and not just the atoms within the sphere:

byres S1


On 4/3/19 4:21 PM, Steiner, Roberto wrote:

pymol?

example
S1 around 12.3  a.  Atoms with centers within 12.3 Angstroms of 
the center of any atom in S1

Best wishes
Roberto

On 3 Apr 2019, at 15:18, Stephen Cusack 
mailto:cus...@embl.fr>> wrote:


Dear All,

  I am looking for an accessible programme that allows selection of 
atoms from a PDB file


within a sphere of inputted radius from a central atom.

Thanks for any help,

Stephen Cusack

--

**
Dr. Stephen Cusack, FRS
Head of Grenoble Outstation of the European Molecular Biology 
Laboratory (EMBL)
Group leader in structural biology of protein-RNA complexes and viral 
proteins

**

Email: cus...@embl.fr
Website: 
https://eur03.safelinks.protection.outlook.com/?url=http%3A%2F%2Fwww.embl.frdata=01%7C01%7Croberto.steiner%40KCL.AC.UK%7C741d46af65624537240e08d6b83f5ac5%7C8370cf1416f34c16b83c724071654356%7C0sdata=he7%2BNc8OllNF1sHbwnp6VcqkEbXUHohZNp9UgrhHMUc%3Dreserved=0

Tel: (33) 4 76 20 7238Secretary (33) 4 76 20 7123
Fax:(33) 4 76 20 7199
Postal address:   EMBL Grenoble Outstation, 71 Avenue des Martyrs, CS 
90181, 38042 Grenoble Cedex 9, France
Delivery address: EMBL Grenoble Outstation, Polygone Scientifique, 71 
Avenue des Martyrs, 38000 Grenoble, France

**



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King's College London

roberto.stei...@kcl.ac.uk
Phone 0044 20 78488216
Fax0044 20 78486435

Room 3.10A
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Re: [ccp4bb] select from a pdb file a sphere of atoms

2019-04-03 Thread Tristan Croll


In ChimeraX:

ctrl-click to select your atom, then in the command line below:

select zone sel 

Best regards,

Tristan

On 2019-04-03 15:18, Stephen Cusack wrote:

Dear All,

 I am looking for an accessible programme that allows selection of
atoms from a PDB file

within a sphere of inputted radius from a central atom.

Thanks for any help,

Stephen Cusack

--

**
Dr. Stephen Cusack, FRS
Head of Grenoble Outstation of the European Molecular Biology 
Laboratory (EMBL)
Group leader in structural biology of protein-RNA complexes and viral 
proteins

**

Email:  cus...@embl.fr
Website: http://www.embl.fr
Tel:(33) 4 76 20 7238Secretary (33) 4 76 20 7123
Fax:(33) 4 76 20 7199
Postal address:   EMBL Grenoble Outstation, 71 Avenue des Martyrs, CS
90181, 38042 Grenoble Cedex 9, France
Delivery address: EMBL Grenoble Outstation, Polygone Scientifique, 71
Avenue des Martyrs, 38000 Grenoble, France
**



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Re: [ccp4bb] Ramachandran outliers

2019-03-08 Thread Tristan Croll

Dear Tereza,

First, a shameless plug for ISOLDE (https://isolde.cimr.cam.ac.uk). It’s built 
specifically for working with models around your resolution.

Other than that, I’d suggest having a close look at the corresponding sites in 
your high-resolution reference models as a sanity check. Remember: just because 
a model is built into high-resolution data doesn’t mean it’s completely correct 
- mistakes happen at all resolutions. Conversely, some outliers are real - if 
it’s an outlier in the reference model *and* well-supported by the 
high-resolution density, you can tick it off your list of things to worry about.

Best regards,

Tristan
 

> On 8 Mar 2019, at 09:08, Tereza Skalova  wrote:
> 
> Dear all,
> 
> I have structure at 3.3A resolution and I have ca. 35 Ramachandran outliers.
> Do you have any idea how to reduce the number?
> I refine in Refmac, using h-bond based Prosmart restraints based on PDB 
> structures (identical molecules with high resolution) and I use NCS, medium 
> between AB (protein 1) and loose between CDE (protein 2). I use overall 
> B-factor and 8 TLS groups.
> Is it possible to optimize Ramachandran plot directly in Refmac?
> 
> Thank you
> 
> Tereza Skalova
> 
> 
> 
> 
> 
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Re: [ccp4bb] 2019 mid-range & high-end LINUX laptops for structural biology

2019-02-13 Thread Tristan Croll

I've been really happy with the performance of my laptop (a 2016-era 
Asus ROG Strix GL502vs) for MD work. It's a gaming model (GTX 1070 
rather than Quadro, and an i7 CPU), but it's solid and seriously 
powerful. I dual-boot Fedora and Windows (I initially tried with Ubuntu, 
but its setup crashed in the middle of rearranging the hard drive 
partitions leaving me too scared to try again). It works nicely enough, 
other than the occasional issue with kernel updates breaking the Nvidia 
driver and/or GRUB.


On 2019-02-13 14:09, Domen Zafred wrote:

Dear CCP4 community,

My 8-year-old Dell Inspiron with Ubuntu 10 is waving goodbye and I
would ask for some help on this bulletin as buying/setting up a new
workstation is common trouble in our community.

Has anyone recently set up a laptop for structural work with all the
MD, docking, etc. _in silico _simulations? Is dual boot Linux/Win
still a thing, or do you rather run Windows virtually (which VM
software)? Are there any known issues with Nvidia Quadro P series GPUs
or Intel i5 processors? Has anyone tried Xeon/ECC on a laptop? Does
Ubuntu LTS still win the hearts of non-enthusiasts?

Any successfully tested laptops (with names and/or listed
configuration) are very welcome and any answers that include fruits or
hamburger names may be kindly avoided :)

Thanks a lot,

Domen

-

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Re: [ccp4bb] List of obsolete residue IDs

2019-02-11 Thread Tristan Croll

It turns out the field I was looking for is 
_chem_comp.pdbx_release_status. Thank you very much to those who pointed 
me to this!


On 2019-02-08 22:31, Tristan Croll wrote:

Hi all,

I’m trying to find an authoritative list of all obsolete residue IDs
in the CCD, but I’m coming up blank. Ligand Expo will tell me if a
given ID is obsolete, but where is this information stored?
Components.cif includes an “Obsoleted” date for a few dozen residues,
but that doesn’t cover all of them. Is there another entry in the .cif
that I’m missing? Any help would be much appreciated.

Thanks,

Tristan



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[ccp4bb] List of obsolete residue IDs

2019-02-08 Thread Tristan Croll

Hi all,

I’m trying to find an authoritative list of all obsolete residue IDs in the 
CCD, but I’m coming up blank. Ligand Expo will tell me if a given ID is 
obsolete, but where is this information stored? Components.cif includes an 
“Obsoleted” date for a few dozen residues, but that doesn’t cover all of them. 
Is there another entry in the .cif that I’m missing? Any help would be much 
appreciated.

Thanks,

Tristan



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[ccp4bb] ISOLDE 1.0b2 release

2019-01-22 Thread Tristan Croll


Dear structural biology community,

I am very happy to announce that ISOLDE 1.0b2 is now available. All the 
details including documentation are available on the new ISOLDE website 
at https://isolde.cimr.cam.ac.uk, but the following is a short list of 
new features and improvements since ISOLDE 1.0b1:


- added support for Windows. ISOLDE is now available for all three major 
operating systems.
- many usability and stability improvements. In particular, ISOLDE is 
now much more helpful when your starting model is not yet quite 
simulation-ready.
- No more need to delete unparameterised residues from your model. You 
can now choose to have ISOLDE simply ignore them for simulation 
purposes.
- Real-time smoothing of the trajectory visualisation to reduce thermal 
jitter.
- Direct handling of real-space maps (no need to convert them to 
structure factors first).
- Live recalculation of structure factors and crystallographic maps from 
F/sigF data as model coordinates change.


I do hope you find it useful.

Best regards,

Tristan



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Re: [ccp4bb] help needed with a rabbit-head-shaped blob

2018-11-03 Thread Tristan Croll

In all seriousness, it looks like it may be some form of hexose sugar?

 
 
Tristan Croll
Research Fellow
Cambridge Institute for Medical Research
University of Cambridge CB2 0XY
 

 

> On 2 Nov 2018, at 21:14, Deborah Harrus  wrote:
> 
> Dear all,
> 
> I came across an unidentified rabbit-head-shaped blob and would need help for 
> its identification. There are 2 molecules of protein per asymmetric unit but 
> there are some differences between the two chains. The blob is located in 
> between the two chains, and is surrounded by residues Asp, Pro, and Val.
> 
> The protein, a glycosyltransferase, was expressed in E. coli BL21(DE3) and 
> purified on Ni-NTA followed by gel filtration. The purification buffer 
> included Sodium phosphate and NaCl. The crystallization condition had 
> Bis-Tris, ammonium acetate and PEG 2, and glycerol was used as 
> cryoprotectant. From the size, bis-tris was the most probable, but I have 
> tried to fit it and it is not convincing.
> 
> The structure is 1.6 angstrom resolution and this is the only thing left to 
> be done, it's driving me crazy!
> 
> Pictures attached show the face, bottom and top of the head. 2Fo-Fc is at 1.1 
> sigma, Fo-Fc at 3 sigmas.
> 
> Many thanks in advance for your suggestions!
> 
> Regards,
> Deborah.
> 
> 
> =
> Deborah Harrus, PhD.
> 
> University of Oulu / Faculty of Biochemistry and Molecular Medicine
> Aapistie 7 A, 90220 Oulu
> Finland
> 
> office: F123B
> email: deborah.har...@oulu.fi
> phone: +358 50 3502387 / +358 44 2386351
> http://www.oulu.fi/fbmm/node/20603
> =
> 
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1
> 
> 
> 
> 



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[ccp4bb] Introducing ISOLDE

2018-05-01 Thread Tristan Croll


Dear members of the structural biology community,

It is with great pleasure that I announce the release of version 1.0b1 
of ISOLDE, a new, free for academic/nonprofit use, environment for 
interactive rebuilding of macromolecular models against low-medium 
resolution experimental crystallographic or cryo-EM maps.


A paper describing ISOLDE is online at 
http://scripts.iucr.org/cgi-bin/paper?S2059798318002425, and you can 
also get an overview from my demonstration video at 
https://www.youtube.com/watch?v=limaUsNAVL8.


In brief, though, ISOLDE is designed to help overcome the substantial 
challenge involved in building high-quality models into “difficult” maps 
(e.g. those with worse than 3Å resolution) by re imagining the task as 
one akin to interacting with a real, “living” molecule. At its heart is 
the use of interactive molecular dynamics: all atoms feel van der Waals 
and electrostatic interactions with their neighbours as you interact 
with them, avoiding clashes and constantly settling towards their 
nearest low-energy state. In addition to this, ISOLDE helps interpret 
what you're looking at by performing real-time rotamer, Ramachandran and 
peptide plane validation, flagging up potential problems directly on the 
model to give you instant feedback on the results of your actions.


Specific features:
- support for protein, nucleic acid, water and some small molecules/ions 
(with substantially expanded support to come in later versions)
- start an interactive simulation covering anything from a few residues 
to your entire model (depending on your hardware) at the click of a 
button
- “always-on” validation updates every time coordinates change (results 
are shown directly as markup on the model itself, and also accessible 
via more traditional tables/plots)
- Interactively apply peptide plane or cis/trans flips and add/remove 
position, distance, rotamer and secondary structure restraints via a 
simple graphical interface
- Smoothly shift any continuous stretch of backbone forward/backward 
along the chain to easily recover from register shift errors
- Save a checkpoint in a running simulation and return to it with a 
button click, allowing you to experiment with tricky regions without 
fear
- Real-time visualisation of crystal symmetry contacts: when the master 
atoms move, they move.
- One-button masking of maps to cover any arbitrary atom selection – 
particularly useful for diagnosing large-scale problems.
- Extensive GUI and API documentation, available by clicking the “Help” 
button in the ISOLDE GUI window.


ISOLDE is built as a plugin to UCSF ChimeraX 
(http://preview.cgl.ucsf.edu/chimerax/download.html) and is currently 
available for modern Linux distros and MacOS, and can be installed from 
within ChimeraX via its ToolShed (Tools/More Tools...). It should run on 
any Linux version that supports ChimeraX itself. NOTE: due to some 
recent changes in the ChimeraX API, the current build will only work 
with the ChimeraX 0.6 release (not the version 0.7 daily builds).


In terms of hardware, ISOLDE is designed based on the understanding that 
not everybody has access to high-performance computing facilities, so 
its minimum requirements are as modest as possible. The most important 
requirement is a GPU capable of OpenCL 1.2 or CUDA processing. My 
“go-to” machine for performance testing is a 2016-model gaming laptop 
equipped with a Nvidia GTX 1070 GPU – this achieves ~20 coordinate 
updates and >30 graphics updates per second with live validation on the 
included 229-residue demo model, indistinguishable from (or slightly 
faster than) my Xeon workstation. At the other end of the scale, it runs 
at usable speeds on my MacBook Air for simulations of a few dozen 
residues, sufficient for many tasks.


I hope you find it useful, and enjoy using it as much as I've enjoyed 
making it. The MolProbity score to beat on the demo model is 1.02 (after 
refinement in your favourite package – ID is in the PDB header so you 
can download reflections from the wwPDB), and bonus points if you can 
identify the mystery ligand (I can't). If you recognise it as your model 
I apologise – I picked it out at random, I swear!


Best regards,

Tristan

Re: [ccp4bb] improper rotation matrix

2018-03-12 Thread Tristan Croll

Assuming you have a good reason for this, you can apply any arbitrary 
transform fairly straightforwardly in ChimeraX. In the console 
(Tools/General/Shell), and assuming your model is the only one loaded:


m = session.models.list()[0]
import numpy
transform_matrix = numpy.array([[r11, r12, r13, tx], [r21, r22, r23, 
ty], [r31, r32, r33, tz]])

from chimerax.core.geometry import Place
transform = Place(matrix=transform_matrix)
m.atoms.coords = transform.moved(m.atoms.coords)

... which could of course be extended into a script to loop over many 
models/transforms with a bit more work.


Hope this helps,

Tristan

On 2018-03-12 10:24, Phil Evans wrote:

The command ROTATE in pdbset should allow to apply any valid rotation
matrix, ie determinant = +1.0

Anything else will distort the coordinates


On 12 Mar 2018, at 10:13, Franck Coste  
wrote:


Hi all,

I'd like to apply an improper rotation matrix to PDB files but it 
seems it's not allowed in pdbset. Does anyone know a program where I 
could do this ?


Thanks in advance.

Regards,

Franck.
--

Re: [ccp4bb] validating a homlology model

2018-03-02 Thread Tristan Croll


Hi Careina,

This is a little confusing. A homology model *is* a set of coordinates 
(usually provided as a PDB file by most servers/packages I know of). The 
MolProbity site at http://molprobity.biochem.duke.edu/ allows you to 
upload your own PDB file, and in my experience is quite forgiving 
regarding format.


Hope this helps,

Tristan

On 2018-03-02 11:44, Careina Edgooms wrote:

Dear all

What programs are best used for validate homology models? I know of
molprobity but if there are no coordinates I cannot use it. Is there a
way to use such programs with homology models?

Also I wish to use pdbepisa for to charaterise dimer interface but
again for homology model this cannot be done as there is no PDB model.
Does anybody know way to use PISA software on my own model that is not
deposited in PDB?

Thank you in advance
Careina

Re: [ccp4bb] Help with assigning density

2018-01-26 Thread Tristan Croll


Yes it can. See 5x49 (residue 603) for a wonderful example.

On 2018-01-26 16:46, Michal Boniecki wrote:

I have positive density around lysine residue (image). This density is
found in all crystals, and only with this lysine. It is solvent
accessible but again not only this one is surface K residue. Can PEG
wrap around lysine like this? Maybe someone else have seen similar
density and knows what it might be?

Thank You

Re: [ccp4bb] Modelling protein/protein interfaces

2018-01-26 Thread Tristan Croll

I've been quite impressed by ClusPro (https://cluspro.bu.edu) in the 
past. Rather than pure rigid-body docking it makes some (pretty good) 
effort to adjust interacting sidechains into reasonable arrangements. 
The advanced options also allow you to specify attractions and 
repulsions between specific residue pairs so you can apply any 
experimental knowledge you have.


On 2018-01-26 12:34, Thomas Krey wrote:

Dear colleagues,

Is there any appropriate tool for docking an interacting protein to a
relatively large protein-protein interface (>2500 A2). We are facing a
hetero-oligomer for which we approximately know the interfaces as well
as the structures of the individual protomers and would like to model
the hetero-oligomer. I have done some small molecule docking before,
but the size of the interface seem to be prohibitive for using the
same tools in the case of such a protein-protein interaction.

Any help or suggestion would be highly appreciated.

Best wishes

Thomas

Re: [ccp4bb] Help needed to make a clue about ligand

2018-01-09 Thread Tristan Croll

Looks like it could be a G-C dinucleotide?

 
 
Tristan Croll
Research Fellow
Cambridge Institute for Medical Research
University of Cambridge CB2 0XY
 

 

> On 10 Jan 2018, at 04:09, Shankar Prasad Kanaujia <spkanau...@gmail.com> 
> wrote:
> 
> Dear All,
> 
> Wishing you all a very happy and prosperous new year 2018
> 
> We have recently solved a structure which contains a huge density in the 
> active site. We modelled a molecule fitting the electron density (please see 
> the attached figures). However, we could not find any known molecule similar 
> to the fitted molecule either in PDB or PubChem databases. Thus, we are not 
> able to correlate its existence in cell.
> 
> The protein was expressed in E. coli cells. The structure is solved at 1.85 
> Angst. We have not added any molecule like this in any step of the 
> experiments.
> 
> Looking forward to having some ways to figure out this problem.
> 
> With best regards,
> Shankar
> 
> 
> -- 
> Shankar Prasad Kanaujia, Ph.D.
> Associate Professor
> Department of Biosciences and Bioengineering
> Indian Institute of Technology Guwahati
> Guwahati - 781039 Assam, India
> Tele: 0361 258 2228
> Fax:  0361 258 2249
> Email: spkanau...@iitg.ernet.in
> Homepage: http://www.iitg.ernet.in/spkanaujia/
> Google Scholar: https://scholar.google.com/citations?user=Zt4JSNYJ=en
>

Re: [ccp4bb] Asp-Asp pair facing each other at <3.0 A distance

2017-12-29 Thread Tristan Croll

In this case, yes - there’s clearly a metal ion coordinated there. But for the 
record, it is entirely possible (if somewhat rare) to get acidic side chains 
directly contacting each other. Look up “carboxyl-carboxylate pair”. They’re 
typically found at low pH and/or in quite protected environments. Think of it 
as having one carboxyl group protonated and H-bonding to the other, except that 
the proton is more-or-less equally shared. Their most common role is as 
pH-dependent conformational switches - very stably bonded below the pKa, 
wanting nothing to with each other above.

 
 
Tristan Croll
Research Fellow
Cambridge Institute for Medical Research
University of Cambridge CB2 0XY
 

 

> On 29 Dec 2017, at 14:18, Robbie Joosten <robbie_joos...@hotmail.com> wrote:
> 
> Hi Anurag,
>  
> There is a metal in the middle there. You need to figure out which one. Have 
> a look at the crystallization conditions. If can also have a look at 
> anomalous signal.
>  
> Cheers,
> Robbie
>  
> From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Anurag 
> Misra
> Sent: Friday, December 29, 2017 14:55
> To: CCP4BB@JISCMAIL.AC.UK
> Subject: [ccp4bb] Asp-Asp pair facing each other at <3.0 A distance
>  
> Dear all,
> I have solved a protein structure at 2.8 A resolution with Rfree 26%. In this 
> structure catalytic His interacts with non catalytic Asp. This His-Asp pair 
> (Chain A) is facing another identical His-Asp pair (in Chain B) which are 
> related with 2-fold ncs (pics are attached) (The molecule exists as a dimer). 
> There is another His-Asp pair in chain C (not shown in attached pic) which 
> faces to the identical pair related to crystallographic 2-fold. (min distance 
> Asp-Asp 2.58 A, His-His 3.6 A and His1-Asp1/His1-Asp2 ~3.0 A)
> How to explain Asp-Asp head on pairs in close proximity? Is there a role of 
> His in making such pairs. Is there any role of change in pKa to allow such 
> interactions? Kindly suggest a few references where similar Asp-Asp pairs 
> have been seen.
>  
> I appreciate your kind attention.
> Thank you so much for helping me.
> Best regards,
> Anurag
>  
>  
>

Re: [ccp4bb] average B-factors

2017-12-20 Thread Tristan Croll

This is quite straightforward in any package with a reasonable scripting 
interface. In ChimeraX, for example:


-load your model
-make your selection 
(https://www.rbvi.ucsf.edu/chimerax/docs/user/selection.html)

-open the shell (Tools/General/Shell), and in it, type:

from chimerax.core.atomic import selected_atoms
sel = selected_atoms(session)
sel.bfactors.mean()

Hope this helps,

Tristan

On 2017-12-20 15:02, Amir Khan wrote:

Dear all,

Sorry for naive CCP4 question, is there a simple way to calculate
average B-factors (side chain, backbone, all)
for only part of a model, such as a domain, loop, etc…

Thanks,
Amir

Re: [ccp4bb] AW: Re: [ccp4bb] AW: Re: [ccp4bb] coordinate transformation

2017-12-18 Thread Tristan Croll

Assuming you have a good reason for doing that, I'd suggest the best
approach would be to first generate a real-space map covering your
original coordinates, and then apply the transform to that. To transform
a volumetric map, all you're actually transforming is the (x,y,z)
coordinate of its origin and the three vectors defining its axes - but I
must confess I don't know off the top of my head any GUI tools to do
that for you. Chimera can almost certainly do it, if you look through
the manual.

On 2017-12-18 15:37, Smith Liu wrote:

thanks. i may mean something other. for example, if i rotate the pdb
by 30 degree (or 29.5 degree), or i shift the pdb along x-axis by
something for example 0.123*a, then how i move the mtz map
correspondingly for the fitting of mtz into the transformed map?

Smith Liu

邮箱：smith_liu...@163.com

签名由 网易邮箱大师 [4] 定制

在2017年12月18日 21:58，herman.schreu...@sanofi.com 写道：

Dear Smith,

The map extends through the whole crystal. What happens is that the
map is calculated around the atom you clicked on during centering.
So by centering on your transformed pdb, you will sample the same
map at a different position. Just load your transformed pdb and
untransformed mtz and try.

If the transformed pdb does not fit the map, something went wrong
during the transformation of your pdb. If you have applied an origin
shift (is not equal to applying a crystallographic symmetry
operation), you have to recalculate the mtz, e.g. by running another
round of refinement.

I hope this is clear so.

Herman

VON: Smith Liu [mailto:smith_liu...@163.com]
GESENDET: Montag, 18. Dezember 2017 14:52
AN: Schreuder, Herman /DE
BETREFF: [EXTERNAL] Re: [ccp4bb] AW: Re: [ccp4bb] coordinate
transformation

you mean the mtz map will transform simutaneously?

SMITH LIU

邮箱：smith_liu...@163.com

签名由 网易邮箱大师 [1] 定制

在2017年12月18日 21:26，herman.schreu...@sanofi.com 写道：

If you use coot with on the fly map calculation (e.g. you load an
mtz and not a map file), you do not need to transform the map.
Otherwise I would recommend to run one more round of refinement and
produce a new map your usual way. This will also get rid of any
rounding errors due to the transformation.

Best,

Herman

VON: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] IM AUFTRAG
VON Smith Liu
GESENDET: Montag, 18. Dezember 2017 14:16
AN: CCP4BB@JISCMAIL.AC.UK
BETREFF: [EXTERNAL] Re: [ccp4bb] coordinate transformation

Dear All,

If I have a set of PDB with the corresponding density map, after I
transform the PDB based on the suggestion of everybody, is any way
to transform the map so that the map will be fit with the
transformed PDB?

Smith

At 2017-12-18 18:39:34, "Eleanor Dodson"
<176a9d5ebad7-dmarc-requ...@jiscmail.ac.uk> wrote:

I showed you pdbset ..

Find the centre of mass for your assembly.

Move it where you will

pdbset xyzin mow.pdb

end

Find CoM 0.7 1.3 -0.2

Hmm - a little thought - centre at 1 -1 0 say

pdbset yzin now.pdb xyzout changed.pdb

symgen x , y-2, z

end

New CoM 0.7 -0.7 -0.2

Eleanor

On 18 December 2017 at 00:19, Edward A. Berry
wrote:

Neat idea!
And do you have a 1-line command for setting all the coordinates to
1,1,1? or 0.1,0.1,0.1 if I still want it near the origin but biased
toward the inside of the positive-going cell?
eab

On 12/14/2017 07:23 PM, James Holton wrote:

What I usually do for this is make a copy of the PDB file and change
all the atom x-y-z positions to "1.000". Then I use something like
reforigin or my "origins.com [2]" script to shift the original
coordinates via allowed symmetry operations, origin shifts, or
perhaps indexing ambiguities until it is as close as possible to the
"reference", which is at 1,1,1. I use 1,1,1 instead of 0,0,0 because
there are generally at least two symmetry-equivalent places that are
equidistant from the origin. Declaring the reference to be a bit
off-center breaks that ambiguity, and also biases the result toward
having all-positive x,y,z values.

In case it is interesting, my script is here:

http://bl831.als.lbl.gov/~jamesh/scripts/origins.com [3]

You need to have the CCP4 suite set up for it to work. Run it with
no arguments to get instructions.

-James Holton

MAD Scientist

On 12/13/2017 5:50 AM, Kajander, Tommi A wrote:

Hello,

If someone could point this out would be very helpful... Wasnt
there a simple script somewhere that would transfer coordinates
close to origin - if they for some reason are not? Just cant find
anything right away. Sure i have done this before...

Thanks,

Tommi

Links:
--
[1]
https://urldefense.proofpoint.com/v2/url?u=https-3A__mail.163.com_dashi_dlpro.html-3Ffrom-3Dmail88d=DwMGbwc=Dbf9zoswcQ-CRvvI7VX5j3HvibIuT3ZiarcKl5qtMPor=HK-CY_tL8CLLA93vdywyu3qI70R4H8oHzZyRHMQu1AQm=VsiJhHbT-qp6n3QHdvilZeqY-0tA4mSqlVkx6nStzhMs=0Zu9ZB12d-Kh3exqmmGcW7PZIzCGKTXlXat7Qffx-lke=
[2]

Re: [ccp4bb] new ContaMiner features

2017-11-23 Thread Tristan Croll

Dear Radu,

I think this is a little harsh. Biology is a fabulously messy thing, and very 
prone to doing the unexpected. See the excellent paper by Niedzialkowska et al. 
at https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4815408/#!po=13.6905 for some 
examples. Sometimes unexpected things (which just happen to have a similar size 
to your target) carry through all the purification steps - I remember having 
terrible trouble isolating his-tagged IGF-I (not for crystallization) from Sf9 
lysates due to a cathepsin-like protease that stuck doggedly to the Ni-NTA 
column even under 8M urea, yet co-eluted in imidazole. Even if contaminant 
proteins are barely visible on your SDS-PAGE gel, if they crystallise easily 
and your target doesn’t...  all these things and many others have happened, and 
have undoubtedly driven the occasional poor grad student to the brink of giving 
it all up.

I guess in these days of relatively cheap and ubiquitous mass spec it may make 
sense to sacrifice a crystal to trypsin digest and MS/MS sequencing just for 
peace of mind, but in the average case I think that’s likely to be overkill. 
Shooting crystals at a synchrotron is now very routine, so I think it makes 
perfect sense to provide a computational check for the (hopefully rare) 
surprise case.

Best regards,

Tristan
 
 
Tristan Croll
Research Fellow
Cambridge Institute for Medical Research
University of Cambridge CB2 0XY
 

 

> On 23 Nov 2017, at 19:35, r...@mrc-lmb.cam.ac.uk wrote:
> 
> Dear Stefan,
> 
> Just a couple of thoughts:
> 
> - first of all I think that Gerard is absolutely right, it would have been
> nice to raise such issues first with the developers. In my experience,
> Staraniso does a fantastic job if used correctly.
> 
> - but if you're OK with public trials, may I ask: why on Earth would anybody
> need ContaMiner? Are you trying to offer some sort of computational cure for
> sloppy biochemistry? There is zero point in crystallizing crap samples, sorry
> to say this. In my 17 or so years in Strubi I've never heard of anybody
> crystallizing a "contaminant", being it a purification tag or whatever.
> 
> I suppose this might have happened to somebody you know, hence the motivation
> to spend time on the bizarre ContaMiner. Which is a pity, a silly outcome
> would only teach people to do their job (or train their robots) properly.
> 
> Best wishes,
> 
> Radu
> 
> -- 
> Radu Aricescu
> MRC Laboratory of Molecular Biology
> Francis Crick Avenue
> Cambridge Biomedical Campus
> Cambridge CB2 0QH, U.K.
> tel: +44-(0)1223-267049
> fax: +44-(0)1223-268305
> www: http://www2.mrc-lmb.cam.ac.uk/group-leaders/a-to-g/radu-aricescu
> 
>> Dear Stefan,
>> 
>> Regarding your final paragraph: your server carries a warning
>> with the exact wording:
>> 
>> "Submitting StarAniso files can give you suspicious results. Use
>> with care!"
>> 
>> It seems rather regrettable that you are posting such a public
>> warning without ever having contacted the STARANISO developers about
>> your observations, nor giving any information about what you call
>> "suspicious" or what the "care" you recommend would consist of.
>> 
>> We have taken a great deal of care ourselves in developing the
>> program and offering it to the community through a server, and the
>> least we would have expected is that any pattern of "suspicious"
>> results would be referred to us so that we could investigate them.
>> There may be some assumptions made in MoRDa that we are not aware of,
>> that might be incompatible with assumptions made in STARANISO - who
>> knows? Or it could be that some particularly badly collected datasets
>> are made to look worse after their anisotropy analysis.
>> 
>> Could we discuss your observations, and what it is exactly that
>> you call "suspicious", before they end up being referred to in such an
>> uninformative manner as some sort of "Government Health Warning"?
>> 
>> I think that would be nice :-) and we would be only too keen to
>> take whatever extra "care" is needed ourselves. We would all learn
>> something.
>> 
>> 
>> With best wishes,
>> 
>>  Gerard.
>> 
>> (on behalf of the STARANISO developers)
>> 
>> --
>>> On Thu, Nov 23, 2017 at 05:02:39PM +0100, Stefan Arold wrote:
>>> Dear Community,
>>> 
>>> A quick message to announce the following two new features on our
>>> ContaMiner web server for the automated detection of unwantedly
>>> crystallised contaminants (
>>> https://strube.cbrc.kaust.edu.sa/contaminer/submit)
>

Re: [ccp4bb] What is the easiest/cheapest way to obtain a library of proline derivatives?

2017-11-13 Thread Tristan Croll

I take it you mean for some semisynthetic approach? One solution that springs 
to mind would be to get one or both of the azide/alkyne “click” proline 
derivatives (e.g. 
http://www.sigmaaldrich.com/catalog/product/aldrich/713643?lang=en=GB or 
http://www.sigmaaldrich.com/catalog/product/aldrich/711977?lang=en=GB). 
Depending on your situation it may be feasible to do a single bulk synthesis 
then click on a range of different groups after the fact.

Tristan Croll
Research Fellow
Cambridge Institute for Medical Research
University of Cambridge CB2 0XY

> On 13 Nov 2017, at 21:15, Jan van Agthoven <janc...@gmail.com> wrote:
> 
> Sorry for being a bit off-topic, but I’m looking to transform the catalytic 
> properties of a protein I’m working on by adding an aromatic moiety on one of 
> its prolines. However none of the proline derivatives I find commercially are 
> really interesting. Does anyone know what would be the cheapest way to obtain 
> customized proline derivatives? Most companies ask for more then $3000 per 
> compound (about half a mg).

Re: [ccp4bb] How to deal with the bad omega angles?

2017-10-10 Thread Tristan Croll

Relying on refinement to fix cis peptide bonds for you is unlikely to 
end well. It looks to me like you really need to spend some time 
investigating and manually rebuilding these first.


On 2017-10-10 13:52, 师扬 wrote:

Dear all,
I am refining a model based on a 4.3A EM density map,and there are
some cis-peptides in the beginning model.
By using phenix.real_space_refine with a very low cis-peptide
threshold (0), all the cis-peptide become to the twisted.

The start Omega angle:
 cis-proline: 31.63 %
 twisted proline: 0.00 %
 cis-general: 11.11 %
 twisted-general: 0.05 %
The final Omega angle:
 cis-proline: 0.00 %
 twisted proline: 27.55 %
 cis-general: 0.00 %
 twisted-general: 6.04 %

My questions are:
1) What is the twisted peptide?
2) Is the amount acceptable at the current resolution?
2) How to refine it?

Thanks in advance!
Yang Shi

【网易自营|30天无忧退货】仅售同款价1/4！MUJI制造商“2017秋冬舒适家居拖鞋系列”限时仅34.9元>>
[1]

Links:
--
[1] 
http://you.163.com/item/detail?id=1165011=web_gg_mail_jiaobiao_9

Re: [ccp4bb] Biotin binding protein

2017-09-24 Thread Tristan Croll

You could try getting in touch with the authors of this: 
https://www.ncbi.nlm.nih.gov/m/pubmed/26833545/.

 
 
Tristan Croll
Research Fellow
Cambridge Institute for Medical Research
University of Cambridge CB2 0XY
 

 

> On 23 Sep 2017, at 19:05, Reza Khayat <rkha...@ccny.cuny.edu> wrote:
> 
> Hi,
> 
> Sorry for the non crystallography question. Is anyone aware of monomeric 
> proteins (<20kDa) that can bind to biotin? We need this for a couple of 
> different projects where biotin covalently modifies a ligand of interest. 
> We'd like to complex the biotin to a protein Thanks.
> 
> Best wishes,
> Reza
> 
> Reza Khayat, PhD
> Assistant Professor
> City College of New York
> Department of Chemistry
> New York, NY 10031

Re: [ccp4bb] Unbranched polysaccharide chain

2017-08-30 Thread Tristan Croll

Try http://www.glycosciences.de/modeling/sweet2/doc/index.php

Cheers,

Tristan

 
 
Tristan Croll
Research Fellow
Cambridge Institute for Medical Research
University of Cambridge CB2 0XY
 

 

> On 30 Aug 2017, at 19:56, Reza Khayat <rkha...@ccny.cuny.edu> wrote:
> 
> Hi,
>  
> Is there a server/program that can generate a chemical picture/PDB from the 
> sequence of a polysaccharide and the type of connections? I’m not so 
> interested in the minimized structure at this point. Thanks.
>  
> Best wishes,
> Reza
>  
> Reza Khayat, PhD
> Assistant Professor
> Department of Chemistry
> City College of New York
> 85 Saint Nicholas Terrace, CDI 2.318
> New York, NY 10031
> http://www.khayatlab.org/
> 212-650-6070
>

Re: [ccp4bb] RMSD between superposed structures without moving

2017-08-28 Thread Tristan Croll

I should learn not to post while distracted. That last line was both 
over-engineered, and wrong. What you want is:


rmsd = sum(numpy.linalg.norm(xyz1-xyz2, axis=1))/len(xyz1)

On 2017-08-28 14:32, Tristan Croll wrote:

In this case calculating the rmsd is easy:

- get the coordinates of each structure as n x 3 numpy arrays. The
Pymol commands for this should look like:

xyz1 = cmd.get_coords('sele1', 1)
xyz2 = cmd.get_coords('sele2', 1)

Then,

rmsd = numpy.linalg.norm(numpy.sqrt((xyz1-xyz2)**2), axis=1)

Tristan Croll
Research Fellow
Cambridge Institute for Medical Research
University of Cambridge CB2 0XY

On 28 Aug 2017, at 10:04, Johannes Sommerkamp
<155b9e78396e-dmarc-requ...@jiscmail.ac.uk> wrote:


Thanks a lot for your answers and the PyMOL mailing list hint. I
didnt had in mind this list.

I read the Pymol Wiki. The commands

align moving, target, cycles=0, transform=0

align moving, target, cycles=0

give identical values for RMSD. So, the only difference is, that
the moving structure is not moved in the graphical output.
Additionally the RMSD with the argument cycles=0 can't be the RMSD
before any movement because the values differ for the super and the
align command. I think its just without refinement.
Since the two structure I want to compare are already aligned based
on the central beta sheet CA atoms, I want to calculate the RMSD
without any movement.

Regards
Johannes

On 27/08/17 19:18, Folmer Fredslund wrote:
Hi Johannes,

Did you read the PymoWIKI entry on the align command?

https://pymolwiki.org/index.php/Align#RMSD [1]

I think this should give you what you want within PyMOL.

Btw, there is a nice dedicated PyMOL mailing list
https://pymolwiki.org/index.php/PyMOL_mailing_list [2]
It is rather low traffic, but the replies are generally from the
developers or very knowledgeable users.

Hope this helps,
Folmer Fredslund

On 2017-08-27 13:09, Johannes Sommerkamp wrote:
Hello everybody,
I have superposed two structures based on the central beta-sheet CA
atoms with the "super" command in Pymol.
Now, I want to calculate the RMSD between ALL atoms or ALL CA atoms
without moving the structures again. The rms_cur command in Pymol
would do that, but only works if all atom identifiers match. Adding
"transform=0" to the super, oder align command still does the
alignment and moves the structure but does not show the movement.

Is there an easy way to just calculate the all atom RMSD between
two already superposed structures in pymol or any other programm?

Thanks in advance!
Johannes


--
Johannes Sommerkamp
Ruhr-Universität Bochum
AG Röntgenstrukturanalyse an Proteinen, LS Biophysik, ND04/396
Universitätsstraße 150
44801 Bochum
Tel: +49-(0)234/32-25754


Links:
--
[1] https://pymolwiki.org/index.php/Align#RMSD
[2] https://pymolwiki.org/index.php/PyMOL_mailing_list

Re: [ccp4bb] RMSD between superposed structures without moving

2017-08-28 Thread Tristan Croll

In this case calculating the rmsd is easy:

- get the coordinates of each structure as n x 3 numpy arrays. The Pymol 
commands for this should look like:

xyz1 = cmd.get_coords('sele1', 1)
xyz2 = cmd.get_coords('sele2', 1)

Then,

rmsd = numpy.linalg.norm(numpy.sqrt((xyz1-xyz2)**2), axis=1)


 
 
Tristan Croll
Research Fellow
Cambridge Institute for Medical Research
University of Cambridge CB2 0XY
 

 

> On 28 Aug 2017, at 10:04, Johannes Sommerkamp 
> <155b9e78396e-dmarc-requ...@jiscmail.ac.uk> wrote:
> 
> Thanks a lot for your answers and the PyMOL mailing list hint. I didnt had in 
> mind this list.
> 
> I read the Pymol Wiki. The commands 
> align moving, target, cycles=0, transform=0
> 
> align moving, target, cycles=0
>  
> give identical values for RMSD. So, the only difference is, that the moving 
> structure is not moved in the graphical output. Additionally the RMSD with 
> the argument cycles=0 can't be the RMSD before any movement because the 
> values differ for the super and the align command. I think its just without 
> refinement.
> Since the two structure I want to compare are already aligned based on the 
> central beta sheet CA atoms, I want to calculate the   RMSD without any 
> movement. 
> 
> 
> Regards 
> Johannes 
> 
> 
> 
>> On 27/08/17 19:18, Folmer Fredslund wrote:
>> Hi Johannes, 
>> 
>> Did you read the PymoWIKI entry on the align command? 
>> 
>> https://pymolwiki.org/index.php/Align#RMSD 
>> 
>> I think this should give you what you want within PyMOL. 
>> 
>> Btw, there is a nice dedicated PyMOL mailing list 
>> https://pymolwiki.org/index.php/PyMOL_mailing_list 
>> It is rather low traffic, but the replies are generally from the developers 
>> or very knowledgeable users. 
>> 
>> Hope this helps, 
>> Folmer Fredslund 
>> 
>>> On 2017-08-27 13:09, Johannes Sommerkamp wrote: 
>>> Hello everybody, 
>>> I have superposed two structures based on the central beta-sheet CA atoms 
>>> with the "super" command in Pymol. 
>>> Now, I want to calculate the RMSD between ALL atoms or ALL CA atoms without 
>>> moving the structures again. The rms_cur command in Pymol would do that, 
>>> but only works if all atom identifiers match. Adding "transform=0" to the 
>>> super, oder align command still does the alignment and moves the structure 
>>> but does not show the movement. 
>>> 
>>> Is there an easy way to just calculate the all atom RMSD between two 
>>> already superposed structures in pymol or any other programm? 
>>> 
>>> Thanks in advance! 
>>> Johannes 
>>> 
> 
> -- 
> Johannes Sommerkamp
> Ruhr-Universität Bochum
> AG Röntgenstrukturanalyse an Proteinen, LS Biophysik, ND04/396
> Universitätsstraße 150
> 44801 Bochum
> Tel: +49-(0)234/32-25754

Re: [ccp4bb] positive density near asparagine

2017-08-19 Thread Tristan Croll

If I've got the site right (the equivalent to Asn833 in your image would 
be Asn858 in 3rjo?) then the first thing I'd be checking is if you have 
an unmodelled tail or loop that could be interacting here across a 
symmetry interface. The dramatic kink in the helix there looks like an 
asparagine, glutamine or serine sidechain could stably H-bond to the 
backbone in a conformation reasonably consistent with that three-pronged 
density. You also have partially-unmodelled lysine residues at the 834 
and 831 positions, and quite a few other positive residues in the 
vicinity. I'd be looking for candidates with negative charge. Could the 
spheroidal blob near Asn833 be a sulfate bridging it and Lys834?


On 2017-08-18 22:17, Maben, Zachary wrote:

Hello,

I am refining a few datasets of my protein of interest (ERAP1) bound
to various inhibitors, and I repeatedly observe a positive density
peak near an asparagine. I first thought this might be a glycan, but
this site does not fit the canonical N-linked glycosylation motif
(this site sequence is NKL). Previous crystal structures of this
protein do not identify any ligand at this position.

https://www.dropbox.com/sh/2wdax5uqlb1f4m3/AABQlGK2_mgibtqnm61E-7o0a?dl=0
[1]

 [1]

 Asn833 positive density [1]
 www.dropbox.com
 Shared with Dropbox

I would appreciate any suggestions of what this density might be.
Thank you for your help.

Best,

Zach Maben

Stern Lab

Pathology Department

University of Massachusetts Medical School

 [1]


Links:
--
[1] 
https://www.dropbox.com/sh/2wdax5uqlb1f4m3/AABQlGK2_mgibtqnm61E-7o0a?dl=0

Re: [ccp4bb] Stable Refinement as Low(ish) resolution

2017-07-13 Thread Tristan Croll

*Ahem*

*ISOLDE

 
 
Tristan Croll
Research Fellow
Cambridge Institute for Medical Research
University of Cambridge CB2 0XY
 

 

> On 13 Jul 2017, at 08:02, Tristan Croll <ti...@cam.ac.uk> wrote:
> 
> This is more-or-less exactly the task I'm building ISODE for. It's early 
> days, still quite rough around the edges and by no means complete, but if you 
> have a machine with a decent GPU running a modern Linux distro, then it's 
> easily available as a plugin to ChimeraX after installing the latest daily 
> build from https://www.rbvi.ucsf.edu/chimerax/download.html. Think of it as 
> an interactive real-space refinement environment, and you won't be far off. 
> This video is getting a little dated now, but should give you some idea of 
> what it can do: 
> https://drive.google.com/open?id=0B6uMjfjuw4k8aUdyTEJWeEJ1UnM. I'm currently 
> working on a longer, narrated demo/tutorial video showing all the latest 
> features (interactive position/rotamer/secondary structure assignment, a very 
> fun tool for shifting stretches in register, ...).
> 
> There are some caveats:
> - It doesn't yet do any building (adding or removing of atoms/residues)
> - It requires all residues to be complete including hydrogens
> - Currently only protein, nucleic acid and metal ions are supported (although 
> the metal ions need more work to be properly useful)
> 
> Let me know if it's of interest, and we can talk more offline.
> 
> Cheers,
> 
> Tristan
> 
>> On 2017-07-13 00:17, Rhys Grinter wrote:
>> Dear All,
>> I'm currently in the process of refining a low(ish) resolution
>> structure at 3.2 Ang, with a fair level of anisotropy. I processed the
>> data through the anisotropy server
>> (https://services.mbi.ucla.edu/anisoscale/ [1]), which elliptically
>> truncated the data to 4.0, 3.8 and 3.2 Ang. This really improved the
>> maps and allowed me to trace the majority of the chain and build most
>> side chains.
>> The R-factors are reasonable (0.29 work and 0.35 free respectively).
>> but I'm having trouble with over fitting in refinement as I continue
>> to refine. What parameters/restraints would the community generally
>> use when refining this kind of structure? Additionally Refmac doesn't
>> seem to read the structure factors from the anisotropy server output
>> file properly, giving vastly inflated R values and strange looking
>> maps.
>> Cheers,
>> Rhys
>> --
>> Dr Rhys Grinter
>> Sir Henry Wellcome Fellow
>> Monash University
>> +61 (0)3 9902 9213
>> +61 (0)403 896 767
>> Links:
>> --
>> [1] https://services.mbi.ucla.edu/anisoscale/

Re: [ccp4bb] Stable Refinement as Low(ish) resolution

2017-07-13 Thread Tristan Croll

This is more-or-less exactly the task I'm building ISODE for. It's early 
days, still quite rough around the edges and by no means complete, but 
if you have a machine with a decent GPU running a modern Linux distro, 
then it's easily available as a plugin to ChimeraX after installing the 
latest daily build from 
https://www.rbvi.ucsf.edu/chimerax/download.html. Think of it as an 
interactive real-space refinement environment, and you won't be far off. 
This video is getting a little dated now, but should give you some idea 
of what it can do: 
https://drive.google.com/open?id=0B6uMjfjuw4k8aUdyTEJWeEJ1UnM. I'm 
currently working on a longer, narrated demo/tutorial video showing all 
the latest features (interactive position/rotamer/secondary structure 
assignment, a very fun tool for shifting stretches in register, ...).


There are some caveats:
- It doesn't yet do any building (adding or removing of atoms/residues)
- It requires all residues to be complete including hydrogens
- Currently only protein, nucleic acid and metal ions are supported 
(although the metal ions need more work to be properly useful)


Let me know if it's of interest, and we can talk more offline.

Cheers,

Tristan

On 2017-07-13 00:17, Rhys Grinter wrote:

Dear All,

I'm currently in the process of refining a low(ish) resolution
structure at 3.2 Ang, with a fair level of anisotropy. I processed the
data through the anisotropy server
(https://services.mbi.ucla.edu/anisoscale/ [1]), which elliptically
truncated the data to 4.0, 3.8 and 3.2 Ang. This really improved the
maps and allowed me to trace the majority of the chain and build most
side chains.

The R-factors are reasonable (0.29 work and 0.35 free respectively).
but I'm having trouble with over fitting in refinement as I continue
to refine. What parameters/restraints would the community generally
use when refining this kind of structure? Additionally Refmac doesn't
seem to read the structure factors from the anisotropy server output
file properly, giving vastly inflated R values and strange looking
maps.

Cheers,

Rhys

--

Dr Rhys Grinter
Sir Henry Wellcome Fellow
Monash University
+61 (0)3 9902 9213
+61 (0)403 896 767

Links:
--
[1] https://services.mbi.ucla.edu/anisoscale/

Re: [ccp4bb] Correcting 3-letter codes based on protonation states in a PDB file

2017-06-29 Thread Tristan Croll

This can be done in a few lines of script in any structural biology 
package that provides a Python (or other) shell. Here's how I'd do it in 
ChimeraX, for example:


Assuming your model is the only one loaded and atom names are all 
standard:


m = session.models.list()[0]
histidines = m.residues.filter(m.residues.names == 'HIS')
for his in histidines:
names = his.atoms.names
he2 = 'HE2' in names
hd1 = 'HD1' in names
if hd1 and not he2:
his.name = 'HID'
elif he2 and not hd1:
his.name = 'HIE'
elif hd1 and he2:
his.name = 'HIP'
else:
raise RuntimeError('HIS {}:{} is missing both 
hydrogens!'.format(

h.chain_id, h.number))

Cheers,

Tristan

On 2017-06-29 07:09, Briggs, David C wrote:

I believe the ProPka or Pdb2pqr webservers can do this.

 ProPka.org

 http:// [1]nbcr [1]-222.ucsd.edu/pdb2pqr_2.0.0/ [1]

 HTH,

 Dave

 --

 Dr David C Briggs

 Hohenester Lab

 Department of Life Sciences

 Imperial College London

 UK

 http://about.me/david_briggs [2]

-

FROM: CCP4 bulletin board  on behalf of
Sampson, Jared 
 SENT: Wednesday, June 28, 2017 11:34:05 PM
 TO: CCP4BB@JISCMAIL.AC.UK
 SUBJECT: [ccp4bb] Correcting 3-letter codes based on protonation
states in a PDB file

Dear all -

 I'm working with a PDB file with explicit hydrogens where many of the
histidines are in protonated form due to crystallization at low pH.
Unfortunately, although the additional protons are present in the
model for the positively charged histidines, the residues in question
are indicated in both the SEQRES and the ATOM records as 3-letter code
`HIS` regardless of protonation state (i.e. instead of `HIP` for
positively charged, and `HID` or `HIE` for the neutral tautomers).

 Are there existing tools available to determine the proper 3-letter
residue code for titratable amino acid residues based on which
hydrogens are present, and output a corrected PDB file?

 Thank you in advance for your suggestions.

 Cheers,

 Jared Sampson
 Ph.D. Candidate
 Columbia University

Links:
--
[1] http://nbcr-222.ucsd.edu/pdb2pqr_2.0.0/
[2] http://about.me/david_briggs

Re: [ccp4bb] Incorrect Structure in the PDB

2017-06-27 Thread Tristan Croll

Note that in most cases a structure may only be obsoleted with the written 
agreement of a senior author. From experience it's by far the best approach to 
work together with the original authors on a correction, in any case. Goodwill 
is an important thing.

If you wish, you can go it alone and deposit a new structure based on data from 
an existing deposition, but it won't obsolete the old structure and I believe 
it will only be accepted if accompanied by a peer-reviewed publication.

 Hope this helps,

Tristan
 
Tristan Croll
Research Fellow
Cambridge Institute for Medical Research
University of Cambridge CB2 0XY
 

 

> On 27 Jun 2017, at 07:50, Vellieux Frédéric <frederic.velli...@ibt.cas.cz> 
> wrote:
> 
> Hello,
> 
> I think it makes sense to have a look at the policy of the PDB concerning 
> obsoleting structures:
> 
> The publication is retracted. The associated PDB entry will be obsoleted if 
> requested by the journal. If a request has not been received, the wwPDB will 
> do its best to contact the depositor and co-authors, (former) PIs, journal 
> editors, etc. when made aware of the retraction. If the reason(s) for 
> retraction were such that the associated PDB entry needs to be made obsolete, 
> the wwPDB will obsolete the entry. The citation in the obsoleted entry is the 
> published journal retraction.
> The structure is incorrect, and the entry author obsoletes the entry. The 
> entry must contain a statement as to the reason for obsoleting the structure.
> A third-party (such as the employer) requests that the entry is obsoleted 
> (e.g., in case of malfeasance). The citation in the obsoleted entry must be a 
> published explanation and retraction in a peer-reviewed journal.
>  
> Source: https://www.wwpdb.org/documentation/policy
> 
> Cheers,
> 
> Fred.
>  
> From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Trevor 
> Sewell
> Sent: Tuesday, June 27, 2017 8:35 AM
> To: CCP4BB@JISCMAIL.AC.UK
> Subject: [ccp4bb] Incorrect Structure in the PDB
>  
>  
> I have come across a key paper in my field that describes an enzyme 
> mechanism. Their work is based on a deposited structure – by other authors - 
> that is incorrectly interpreted.
>  
> Is there a process for removing a demonstrably wrong structure (deposited by 
> others) from the PDB and replacing it with a correctly interpreted structure 
> based on the original data? Or is there an alternative, and generally 
> recognized, way of getting the correct structure in the public domain?
>  
> Many thanks for your advice on this matter.
>  
> Trevor Sewell
>  
> Disclaimer - University of Cape Town This e-mail is subject to UCT policies 
> and e-mail disclaimer published on our website at 
> http://www.uct.ac.za/about/policies/emaildisclaimer/ or obtainable from +27 
> 21 650 9111. If this e-mail is not related to the business of UCT, it is sent 
> by the sender in an individual capacity. Please report security incidents or 
> abuse via cs...@uct.ac.za
> - 
> Upozornění: Není-li v této zprávě výslovně uvedeno jinak, má tato E-mailová 
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> nezavazují. Text této zprávy nebo jejích příloh není návrhem na uzavření 
> smlouvy, ani přijetím případného návrhu na uzavření smlouvy, ani jiným 
> právním jednáním směřujícím k uzavření jakékoliv smlouvy a nezakládá 
> předsmluvní odpovědnost Biotechnologického ústavu AV ČR, v. v. i. 
> Disclaimer: If not expressly stated otherwise, this e-mail message (including 
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Re: [ccp4bb] off-topic:fluorescence polarization displacement assay

2017-06-21 Thread Tristan Croll

Looks like positive cooperativity to me. If binding at one site increases 
affinity at the second site, then intermediate concentrations of your 
unlabelled ligand can (seemingly paradoxically) *increase* binding of your 
tracer. At higher levels still, the unlabelled ligand outcompetes the tracer as 
expected. See figure 1 in http://molpharm.aspetjournals.org/content/70/5/1783.

Hope this helps,

Tristan

 
 
Tristan Croll
Research Fellow
Cambridge Institute for Medical Research
University of Cambridge CB2 0XY
 

 

> On 21 Jun 2017, at 20:20, megha abbey <abbey...@gmail.com> wrote:
> 
> Hello All,
> 
> This is an off-topic question. I have some issues regarding Fluorescence 
> Polarization competitive displacement assay and would need some advice. 
> 
> I have developed an in vitro fluorescence polarization based assay using a 
> N-terminus labelled FITC peptide. The peptide is 21 amino acids long and the 
> binding protein is 50 Kda in size. The Kd for interaction is 1 uM. I am 
> further running a competitive displacement assay using the exactly same 
> unlabelled peptide. Here, with increasing concentration of the unlabeled 
> peptide, the polarization signal first increases and then shows a sharp 
> decline. Attached is the raw data file for the above. 
> 
> I believe that if the unlabelled peptide had been aggregating, the 
> polarization signal would increase but not drop. If the binding would have 
> been non-specific, then the unlabelled peptide should not displace at all, 
> but here I see an increase followed by a decrease in the signal. What does 
> this increase and sharp drop in polarization signify and how do I fix this? 
> Please help.
> 
> I have checked the polarization for titration of the unlabelled peptide mixed 
> with fixed conc. of FITC-peptide (no protein added). Here, the polarization 
> signals are the same for the entire range of unlabeled peptide. I have also 
> tried incubating the unlabelled peptide with the protein (for ~15min) first 
> followed by addition of FITC-peptide, but the results are the same.  
> 
> Thank you,
> Megha
>

Re: [ccp4bb] similar unit cell

2017-06-17 Thread Tristan Croll

With those statistics, it seems most probable that these two crystals 
are the same protein. Do your two target proteins share an expression 
system and/or purification protocol that could lead to the same 
contaminant in both? If you have the resolution you could try Arcimboldo 
to get an initial solution and perhaps infer the sequence from the 
density. If you have crystals to spare you could digest one and identify 
it via tandem mass spec. You might also try ContaMiner.


Good luck!

Tristan

On 2017-06-17 09:07, dongxiaofei wrote:

Dear ALL,

I got two kinds of crystals of different proteins ,but there are many
similarities.
The shape of the crystals are similar, the cell parameters are also
similar :
 protein A , 136.12 94.398 89.476 90 125.479 90 , Space group C 1 2 1
and
 protein B , 136.14 94.369 89.115 90 125.495 90 , Space group C 1 2 1.


protein A has a NMR structure ,but Rfree always high above 50% after
molecular replacement , protein B’s Rfree is also above 50% .

So I am wonder if these crystals are the result of debris of proteins
, because the growth of the crystals needs more than half a year . I
am sure the two proteins are different and crystals respectively come
from different proteins

Any insights will be really appreciated.

Thanks

Dong Xiao

Re: [ccp4bb] peroxy-glutamate?

2017-05-09 Thread Tristan Croll

A slightly different wrinkle on the perennial "do we model unresolved 
sidechains" debate, I guess. I would argue that in the case of mutations, tags 
etc. those are things you know were there before you started firing x-rays at 
your sample. In the case of the sidechain decarboxylation we know it's an 
artifact of the data collection method, and we correct for known artifacts in 
other contexts all the time.

 
 
Tristan Croll
Research Fellow
Cambridge Institute for Medical Research
University of Cambridge CB2 0XY
 

 

> On 9 May 2017, at 17:49, Ian Tickle <ianj...@gmail.com> wrote:
> 
> 
> Hi Tristan
> 
> I'm not so sure.  The co-ordinates are the result of the experiment.  How 
> other people choose to interpret those results is their affair.  Taking it to 
> its logical conclusion suppose that we 'damage' the protein by 
> mutating/deleting some residues or adding tags purely for the purpose of 
> getting it to crystallise, do we report the structure of the protein as it is 
> in the crystal, or do we report what it would have been if we hadn't messed 
> with it ?  The choice is clear in that situation.
> 
> Cheers
> 
> -- Ian
> 
>> On 9 May 2017 at 16:45, Tristan Croll <ti...@cam.ac.uk> wrote:
>> Hmm... this is a bit of a philosophical pickle in my mind. Do we want to 
>> model the structure as what it looks like after radiation damage has had its 
>> way with it, or what it must have looked like *before* the damage? I can see 
>> arguments both ways (and can sympathise with the former if you want to make 
>> radiation damage a subject of your manuscript), but this is going to lead to 
>> headaches for people who want to make use of the resulting coordinates to 
>> study the actual biology of your protein. Personally, I'd strongly prefer 
>> the latter approach.
>> 
>> Tristan
>> 
>> 
>>> On 2017-05-09 16:06, Edward A. Berry wrote:
>>>> On 05/09/2017 06:18 AM, Ian Tickle wrote:
>>>> We have seen almost identical density to Ed's for GLU side-chains, with 
>>>> what looks like a linear molecule (yes exactly the size of CO2!) where the 
>>>> carboxylate group would be and absolutely no density for the CG-CD bond.  
>>>> So it's indeed very tempting to say that the CO2 is still there, and 
>>>> presumably making the same H bonds that the carboxylate was making to hold 
>>>> it there.  It would not be hydrated to carbonic acid, according to 
>>>> https://en.wikipedia.org/wiki/Carbonic_acid : "The hydration 
>>>> <https://en.wikipedia.org/wiki/Hydrate> equilibrium constant 
>>>> <https://en.wikipedia.org/wiki/Equilibrium_constant> at 25 °C is called 
>>>> K_h , which in the case of carbonic acid is [H_2 CO_3 ]/[CO_2 ] ≈ 
>>>> 1.7×10^−3 in pure water^[5] 
>>>> <https://en.wikipedia.org/wiki/Carbonic_acid#cite_note-HS-5> and ≈ 
>>>> 1.2×10^−3 in seawater <https://en.wikipedia.org/wiki/Seawater>.^[6] 
>>>> <https://en.wikipedia.org/wiki/Carbonic_acid#cite_note-SB-6> Hence, the 
>>>> majority of the carbon dioxide is not converted into carbo
>>> n
>>> ic
>>>> acid, remaining as CO_2 molecules.".
>>> 
>>> It looks like this ignores subsequent ionization of H2CO3 which would
>>> be quite spontaneous at neutral pH.  However the Wikipedia article
>>> also indicates the equilibrium is quite slow (which makes sense-
>>> otherwise why would carbonic anhydrase exist?) and it would be a great
>>> deal slower in vitreous ice at 100 K. Anyway, I had reached the same
>>> conclusion and have modeled a number of the troublesome glutamates as
>>> decarboxylated with CO2 hovering above. There is a problem that the
>>> remaining CG tends to push the CO2 a little out of the density in some
>>> cases, but not a severe clash and it may work itself out with further
>>> refinement or manual assistance.
>>> eab
>

Re: [ccp4bb] peroxy-glutamate?

2017-05-09 Thread Tristan Croll

Hmm... this is a bit of a philosophical pickle in my mind. Do we want to 
model the structure as what it looks like after radiation damage has had 
its way with it, or what it must have looked like *before* the damage? I 
can see arguments both ways (and can sympathise with the former if you 
want to make radiation damage a subject of your manuscript), but this is 
going to lead to headaches for people who want to make use of the 
resulting coordinates to study the actual biology of your protein. 
Personally, I'd strongly prefer the latter approach.


Tristan

On 2017-05-09 16:06, Edward A. Berry wrote:

On 05/09/2017 06:18 AM, Ian Tickle wrote:
We have seen almost identical density to Ed's for GLU side-chains, 
with what looks like a linear molecule (yes exactly the size of CO2!) 
where the carboxylate group would be and absolutely no density for the 
CG-CD bond.  So it's indeed very tempting to say that the CO2 is still 
there, and presumably making the same H bonds that the carboxylate was 
making to hold it there.  It would not be hydrated to carbonic acid, 
according to https://en.wikipedia.org/wiki/Carbonic_acid : "The 
hydration  equilibrium constant 
 at 25 °C is 
called K_h , which in the case of carbonic acid is [H_2 CO_3 ]/[CO_2 ] 
≈ 1.7×10^−3 in pure water^[5] 
 and ≈ 
1.2×10^−3 in seawater .^[6] 
 Hence, 
the majority of the carbon dioxide is not converted into carbo

n
ic

acid, remaining as CO_2 molecules.".


It looks like this ignores subsequent ionization of H2CO3 which would
be quite spontaneous at neutral pH.  However the Wikipedia article
also indicates the equilibrium is quite slow (which makes sense-
otherwise why would carbonic anhydrase exist?) and it would be a great
deal slower in vitreous ice at 100 K. Anyway, I had reached the same
conclusion and have modeled a number of the troublesome glutamates as
decarboxylated with CO2 hovering above. There is a problem that the
remaining CG tends to push the CO2 a little out of the density in some
cases, but not a severe clash and it may work itself out with further
refinement or manual assistance.
eab

Re: [ccp4bb] peroxy-glutamate?

2017-05-03 Thread Tristan Croll

Peroxyacids are unlikely - they're very reactive/unstable under normal 
conditions. Is it possible your crystal is just at unusually low pH so these 
acids are protonated? That makes the carbon-oxygen bond lengths asymmetric, 
possibly by enough to explain your blobs.

Tristan

 
 
Tristan Croll
Research Fellow
Cambridge Institute for Medical Research
University of Cambridge CB2 0XY
 

 

> On 3 May 2017, at 05:29, Edward A. Berry <ber...@upstate.edu> wrote:
> 
> I'm finishing up refinement of a 1.8A structure (R's 0.17, 0.20) , and among 
> the largest peaks in the difference map are small spherical blobs that seem 
> to be attached (1.46 A here) to carboxylate O's (Figures). Are these likely 
> artifacts? If not, how can I interpret/model them? One idea is that the acid 
> has reacted with peroxide from the PEG to make the (hydro)peroxy-acid. I 
> don't know how stable that would be, and I don't see any peroxyglutamate in 
> Ligand Depot or HIC-Up. Another guess would be acid hydroxamate but I don't 
> know how that would be generated. Methyl ester seems to be ruled out by the 
> proximity of the two water molecules (2.45 and 2.48 A here) suggesting the 
> mystery atom is an H-bond acceptor or donor. However since the occupancy 
> seems to be < 1, the waters may be there only when the atom is not.
> I guess another possibility is there is a lot of motion in the plane of the 
> carboxylate (up and down here) which cannot be modeled by my isotropic 
> B-factors. In some cases the green blobs appear on both sides of the 
> carboxylate (but that could also be alternate conformations of 
> peroxyglutamate).
> 
> The difference map (mFo-DFc, green) is contoured at 3 sigma (.06 e-/A^3). The 
> difference peak is 5.4 sigma (0.1 e/A3).
> The 2mFo-DFc map is contoured at 1.5 sigma (0.1 e/A3). 2mFo-DFc density 
> extends to the difference peak if I contour down at 0.64 sigma (0.04 e/A3, 
> third figure).
> 
> If I put an O atom there, link it with plenty of slack, and refine occupancy, 
> it goes to 1.54 A from the carboxylate O and refines to occupancy 0.35, 
> B-factor 15 (carboxylate O is 30). Now it is reached by 2mFo-DFc density at 
> 1.5 sigma (0.1 e/A3).
> Any suggestions would be welcome.
> eab
> 
> 
>

Re: [ccp4bb] discriminate between solutions from phaser

2017-05-01 Thread Tristan Croll

It seems to me that this could be easily explained by there being a kink near 
the middle of the chain (so the bundle adopts a broad 'V' shape). Then 
fragments up to the length of the "arms" would each give two different but good 
solutions, but fragments longer than that would fall off. Have you tried asking 
Phaser to search for two copies of your 57aa fragment?

 
 
Tristan Croll
Research Fellow
Cambridge Institute for Medical Research
University of Cambridge CB2 0XY
 

 

> On 30 Apr 2017, at 23:26, Dr A.A. Jalan <aa...@cam.ac.uk> wrote:
> 
> Dear all,
> 
> I have a two part question,
> 
> I am trying to solve the structure of a collagen triple helix containing 
> 144aa. Phaser on models containing 21, 30, 39 and so on upto 120aa shows an 
> interesting trend. The TFZ and LLG scores peak at 57aa and then start to fall 
> off significantly. How should one go about explaining this.
> 
> The second question is, phaser yields the following two solutions in the .sol 
> file using 57aa
> 
> #   CLUELESS_57
> SOLU RESOLUTION 1.36573
> 
> SOLU SET RFZ=9.7 TFZ=7.3 PAK=0 LLG=502 TFZ==11.1 LLG=1151 TFZ==16.6 PAK=0 
> LLG=1151 TFZ==16.6
> SOLU HISTORY  RF/TF(8/2:40)RNP(40:4)RNP(4:1)RNP(1:1)
> SOLU SPAC P 1 21 1
> SOLU 6DIM ENSE ensemble1 EULER   86.543   77.471  296.818 FRAC 0.28741 
> -0.00948 0.33176 BFAC -4.77330 #TFZ==16.6 # CLUSTER 2
> SOLU ENSEMBLE ensemble1 VRMS DELTA -0.4975 #RMSD  0.81 #VRMS  0.40
> 
> SOLU SET RFZ=9.3 TFZ=8.7 PAK=0 LLG=494 TFZ==11.3 LLG=1069 TFZ==16.4 PAK=0 
> LLG=1069 TFZ==16.4
> SOLU HISTORY  RF/TF(39/1:33)RNP(33:8)RNP(8:2)RNP(2:2)
> SOLU SPAC P 1 21 1
> SOLU 6DIM ENSE ensemble1 EULER  353.101   26.529  183.602 FRAC 0.12571 
> -0.00662 0.18389 BFAC -4.69482 #TFZ==16.4 # CLUSTER 1
> SOLU ENSEMBLE ensemble1 VRMS DELTA -0.4884 #RMSD  0.81 #VRMS  0.42
> 
> As can be noted, the solutions have fairly different coordinates. How should 
> one go about finding out which one is correct.
> 
> Any inputs and suggestions are greatly appreciated.
> 
> Thank you
> 
> Abhishek
> 
>

Re: [ccp4bb] CH-bond length discrepancies

2017-04-28 Thread Tristan Croll

I believe the reason for the discrepancy here is that MolProbity by default 
places the hydrogens according to the centroid of the electron cloud (most 
relevant to x-ray crystallographic data) rather than the nuclear position. In a 
polar bond like N-H the difference can be quite substantial.

Cheers,

Tristan

 
 
Tristan Croll
Research Fellow
Cambridge Institute for Medical Research
University of Cambridge CB2 0XY
 

 

> On 28 Apr 2017, at 18:33, Bernhard Rupp <hofkristall...@gmail.com> wrote:
> 
> Dear Fellows of the Bond,
>  
> when validating a QM refined homology model with Molprobity, I noticed 
> various 8 sigma deviations in the carbon-hydrogen bond distances.
> Out of curiosity, I then used refmac to calculate riding Hs for the same 
> model, and at least in one instance (N-H backbone) there are
> significant differences between Molprobity and Refmac H bond distances 
> (differences to the QM distances in other
> instances I find interesting, but less relevant for us).
>  
> The riding H vs Molprobity presumably should be consistent, because if we use 
> them in VDW restraints but
> they differ from the validation target, systematic bias will occur. I have no 
> feel how significant that effect
> might be – maybe someone more erudite can comment.
>  
> Examples
>  
> distance  MP   REF QM
> backbone N-H   0.861.011.00  
> phenyl C-H 0.930.931.09
>  
> Best, BR
>  
> PS: If someone has accurate experimental values for CH distances I’d 
> appreciate a link.
> No access to CSD.
>  
> --
> Bernhard Rupp
> Crystallographiae Vindicis Militum Ordo
> http://www.hofkristallamt.org/
> b...@hofkristallamt.org
> +1 925 209 7429
> +43 767 571 0536
> --
> Many plausible ideas vanish
> at the presence of thought
> --
>

Re: [ccp4bb] Sliconizing of cover-slips

2017-04-28 Thread Tristan Croll

Ahh, this brings back memories of a former life, preparing hydrophobic 
coverslips for my surface chemistry experiments. I used chlorotrimethylsilane, 
and what I remember best is that the secret to a good coating is making sure 
your coverslips are utterly clean and dry. I used to clean them by boiling in 
base piranha (concentrated ammonia and hydrogen peroxide - need I say this 
stuff should be treated with extreme respect?) before rinsing thoroughly in 
milli-Q water and baking under vacuum. Then essentially as David said: dump 
them in the silane solution (again, it helps to make sure to keep your solvent 
dry), then fish them out one-by-one, rinse and dump them into a beaker of fresh 
solvent (acetone, I think I used). Then once they're all done, take them out of 
that, dry in a nitrogen stream and store. Painful & fiddly, but you can easily 
do a hundred or so in an afternoon.

Tristan Croll
Research Fellow
Cambridge Institute for Medical Research
University of Cambridge CB2 0XY

> On 27 Apr 2017, at 10:26, Praveen Kumar Tripathi 
> <tripathipraveen.i...@gmail.com> wrote:
> 
> Dear all,
> 
> sorry for off topic question.
> 
> May i know if anybody uses homemade silinization of coverslips for protein 
> crystallization purposes?
> 
> I have purchased Sigmacote SL2-100 ml for silanizing coverslips for hanging 
> drop protein crystallization setup.
> 
> Please share your methods to siliconize coverslip using Sigmacote SL2-100 ml 
> if anybody uses.
> 
> Thanks in advance
> 
> regards
> Praveen
> 
> 
> 
> -- 
> Praveen Kumar Tripathi
> PhD Research Scholar
> Kusuma School of Biological Sciences
> Indian Institute of Technology Delhi-110016
> +91-9873625228

Re: [ccp4bb] Off-topic: Attach His-tagged Protein to Coverslip

2017-04-05 Thread Tristan Croll

One possibility: surface-immobilised forms of the peptide HGGHHG bind strongly 
to His tags (coordinating around zinc rather than nickel). See 
https://www.ncbi.nlm.nih.gov/m/pubmed/15050643/. If you want true permanence, 
you could apply the trick used on BiaCore chips. They use a NTA-functional 
carboxymethyldextran surface. His-tagged proteins are captured as usual, then 
rinsed and made permanent by incubating with some water-soluble carbodiimide to 
link the carboxymethyl groups to surface lysines - the advantage being that all 
the proteins are in essentially the same orientation. Not sure if anyone sells 
coverslip equivalents.

Best regards,

Tristan

Tristan Croll
Research Fellow
Cambridge Institute for Medical Research
University of Cambridge CB2 0XY

> On 5 Apr 2017, at 04:19, Keller, Jacob <kell...@janelia.hhmi.org> wrote:
> 
> Does anyone have a simple way to attach purified his-tagged protein solidly 
> to a coverslip?
>  
> Thanks,
>  
> Jacob Keller
>  
> ***
> Jacob Pearson Keller, PhD
> Research Scientist
> HHMI Janelia Research Campus / Looger lab
> Phone: (571)209-4000 x3159
> Email: kell...@janelia.hhmi.org
> ***
>

Re: [ccp4bb] jhbuild system on Mac

2017-02-16 Thread Tristan Croll


Well, that's nice and straightforward. Thanks!

On 2017-02-16 10:15, Stuart McNicholas wrote:

I set DYLD_LIBRARY_PATH on the command line to point to location of
the svn libraries.

On 16 February 2017 at 10:09, Tristan Croll <ti...@cam.ac.uk> wrote:


Hi all,

While using the ccp4-jhbuild system to rebuild specific components
on the Mac, I'm running into the bug detailed at
https://bugs.launchpad.net/bzr-mac-installers/+bug/1244668 [1]
whenever 'bzr info' is run. The workaround described there no longer
appears to to work for OSX Sierra. Before I go chasing it down the
rabbit hole, I'm just wondering if there is a proper fix?

Thanks,

Tristan




Links:
--
[1] https://bugs.launchpad.net/bzr-mac-installers/+bug/1244668

[ccp4bb] jhbuild system on Mac

2017-02-16 Thread Tristan Croll


Hi all,

While using the ccp4-jhbuild system to rebuild specific components on 
the Mac, I'm running into the bug detailed at 
https://bugs.launchpad.net/bzr-mac-installers/+bug/1244668 whenever 'bzr 
info' is run. The workaround described there no longer appears to to 
work for OSX Sierra. Before I go chasing it down the rabbit hole, I'm 
just wondering if there is a proper fix?


Thanks,

Tristan

Re: [ccp4bb] Unknown electron density blob, pdb convention for partially ordered ligands

2017-01-25 Thread Tristan Croll

I've often wondered about PEG (and, I guess, other synthetic polymers): 
wouldn't it just be better to define the monomer, and then model a chain of 
however many monomers you need?

T

 
 
Tristan Croll
Research Fellow
Cambridge Institute for Medical Research
University of Cambridge CB2 0XY
 

 

> On 25 Jan 2017, at 17:21, Edward A. Berry <ber...@upstate.edu> wrote:
> 
> Uma's use of quotes around "di" suggests a related question about PDB 
> convention. It was my (perhaps not very good) understanding that ligands 
> should be identified by what is actually present in the crystal, and not by 
> what can be modeled. For example endogenous ubiquinone is likely to be UQ50 
> (depending on the species) but most of that 50-carbon side chain is hanging 
> out in the lipid or detergent and completely disordered. Still we should use 
> the ligand identifier for UQ50, even though codes exist for UQ with 5 or 
> 10-carbon side chains that are much better accommodated by the density.
> 
> If that is the case, one should not use the pdb identifier for diethylene 
> glycol (PEG) when PEG4k was the precipitant, unless you believe that the 
> binding site has specifically selected diethylene glycol from an extremely 
> broad range of polymer lengths in the added material.  Using the identifier 
> for a much longer PEG will result in a large number of "missing atoms" listed 
> in the report, but would eliminate the unreasonable assumption that PEG 
> fragment models must always end with a terminal oxygen.
> 
> Even if that is the rule, I would agree that PEGs would be a good place to 
> ignore the rule. Since PEGs have a MW distribution, it is impossible to know 
> exactly what is bound and it may be different in different unit cells. If you 
> are not going to get it right no matter what you put, you might as well put 
> something that fits.
> eab
> 
>> On 01/25/2017 09:51 AM, Uma Gabale wrote:
>> Dear all,
>> Thank you very much for your replies. It is a PEG, a "di"ethylene glycol to 
>> be precise, in most chains.
>> Best regards,
>> Uma.
>> --
>> Uma Gabale, PhD
>> Research Associate
>> Molecular and Cellular Biochemistry
>> Indiana University Bloomington
>>

Re: [ccp4bb] Fwd: [ccp4bb] Remittance advice - Invitation to edit

2016-12-11 Thread Tristan Croll

Phishing spam. Don't click.

T

 
 
Tristan Croll
Research Fellow
Cambridge Institute for Medical Research
University of Cambridge CB2 0XY
 

 

> On 11 Dec 2016, at 17:45, Anindito Sen <andysen.to...@gmail.com> wrote:
> 
> Guys 
> 
> are you all receiving the same??? What on earth is this ?
> 
> Andy
> 
> 
> 
>> Begin forwarded message:
>> 
>> From: Hai-fu Fan <fanha...@gmail.com>
>> Subject: [ccp4bb] Remittance advice - Invitation to edit
>> Date: December 12, 2016 at 2:31:56 AM GMT+9
>> To: CCP4BB@JISCMAIL.AC.UK
>> Reply-To: Hai-fu Fan <fanha...@gmail.com>
>> 
>> Please find attached for your review.
>> Regards.
>> 
>> Hai-fu Fan
> 
>

Re: [ccp4bb] Density for an unknown ligand

2016-11-29 Thread Tristan Croll

Benzopinacol? It's sometimes used as the initiator for radical 
polymerisations, so its presence in plasticware wouldn't be unheard of.


On 2016-11-27 10:33, Wei Liu wrote:

Dear all,

We have recently crystallized a recombinant protein produce from E.
coli, and determined the structure at 1.9 Å. The asymmetric unit
contains two protein monomers. Beyond our expectation, strong Fo - Fc
density is present at a cleft of one subunit, but not in the other.
Density maps are given in the snapshots attached to this email. We
tried to model Tris or Bis-tris propane that were used as the
purification and crystallization buffers, but apparently either of
them poorly fitted in this density. The molecule that can be modeled
here seems more likely to be a ligand comprising 3 or 4 rings with
good planarity, however we did not add any additives in our
crystallization trials. So we think it should be something from E.
coli, which has high affinity to our protein. I wonder if anybody can
figure out which molecule well fits to the electron density.

Best wishes
Wei Liu

Re: [ccp4bb] [ccp4bb] Fwd: [ccp4bb] just out of totally idle curiosity ...

2016-11-10 Thread Tristan Croll

Sorry - didn't realise that was going out to the whole list, and just 
made a liar out of myself. Will bow out now.


On 2016-11-10 10:20, Tristan Croll wrote:

Hi Elton,

I certainly didn't say I agreed with their choice or thought it was a
good one. But as someone who grew up in a low-income, low-employment
environment myself this has the ring of truth to it. I totally agree
with you (and worry just as much) regarding the proliferation of
anti-expert sentiment - but I don't think it's necessarily the biggest
cause of what just happened in this case. It seems to me that a great
many people didn't vote for Trump because of who he was, but *in
spite* of who he was - given the choice between two people who they
perceived as corrupt, malevolent billionaires, they went for the one
who wasn't part of the political machine they see as irrevocably
broken, and at least said some things to indicate that he noticed
them. Essentially a protest vote, just on a country-wide scale. I just
hope the consequences won't be as severe as we all fear.

My personal answer to your final question comes in two parts:

(1) As I understand it, there's at least some indication that it isn't
as bad as it seems (or, more precisely, it's always been at least this
bad, but we just couldn't see it as clearly before). History is
littered with extreme examples of anti-science gaining control at the
highest levels of government (Lysenkoism, witch hunts, creationism,
etc.). At least these days it seems science is mostly on the front
foot (anti-science movements and politicians seem to be mostly
fighting a slowly losing battle against concepts introduced by
scientists, rather than scientists mostly fighting back against
unscientific dogma).

(2) A big part of the solution has to be in communication. Our ability
to communicate our work (and its importance) directly to the public
has never been greater. That "directly" bit is important, because
science journalists and even university PR departments have
historically had a habit of both mangling the science and overblowing
its impact between the scientist and the newsprint. Providing clear,
accessible and, above all, friendly explanations of key work online
can count for a heck of a lot.

Yes, we're certainly facing big problems - but I'm not yet ready to 
despair.


Best regards,

Tristan





On 2016-11-10 09:47, Elton Zeqiraj wrote:

Hi Tristan,

I’m afraid I don’t get the logic in the article you sent. In this
case the farmers who are crying for the attention of the people in
shiny palaces have voted for one who lives in golden mansions.

For me the really worrying thing here is how can so many people be
misled by a false prophet who lies and misbehaves so much. It is a
systemic problem in our society. We never before had so much access to
information, and yet it is so easy to mislead people.

People understood what Trump was and they basically said: “I don’t
care”! We are living in a world where people don’t care about
facts, they just say “I know it to be true”. As scientists whose
job and mission in life is to further and pass knowledge, it is very
serious that we are in this post-truth era. It affects us all when we
talk about climate change, evidence based treatments in our hospitals,
GM debate, etc.

Instead of making this about Trump, I would like to pose a different
question: How are we going to deal with the anti-expert movement that
is now so prominent in our society?

Cheers,
Elton


On Nov 10, 2016, at 8:15 AM, Tristan Croll <ti...@cam.ac.uk> wrote:

In the interests of promoting understanding... the link below is to
an article on what is ostensibly a comedy website and contains a bit
of coarse language, but nevertheless is quite possibly the most
insightful exposition of the situation I've come across. The
two-sentence synopsis: don't think of this as the forces of hate,
fear and ignorance winning. Think of it as a cry for attention from
a large number of people who are seriously struggling and (mostly
correctly) see their problems as being ignored by the system.
Writing them off as ignorant and hateful isn't the answer.

In agreement with various others, this is my first and last post
referencing politics or religion on this forum.



http://www.cracked.com/blog/6-reasons-trumps-rise-that-no-one-talks-about/

[1]

On 2016-11-10 08:00, Marjolein Thunnissen wrote:
Dear Bill,
I fully agree with you, awareness has to be spread and one should
not
ignore politics completely, especially when there are so strong
anti-intellectual (anti-science) statements out there.
best regards
Marjolein
On 10 Nov 2016, at 04:17, William G. Scott <wgsc...@ucsc.edu> wrote:
Dear Edward et al:
I agree we shouldn’t engage in partisan arguments on the CCP4bb.
However, I think it is a mistake, and perhaps a missed opportunity,
to ignore politics completely.
For example, Newt Gingrich is currently in the running for Sec HHS.
He has previously written editorials in the NYT and Wall Str

Re: [ccp4bb] [ccp4bb] Fwd: [ccp4bb] just out of totally idle curiosity ...

2016-11-10 Thread Tristan Croll


Hi Elton,

I certainly didn't say I agreed with their choice or thought it was a 
good one. But as someone who grew up in a low-income, low-employment 
environment myself this has the ring of truth to it. I totally agree 
with you (and worry just as much) regarding the proliferation of 
anti-expert sentiment - but I don't think it's necessarily the biggest 
cause of what just happened in this case. It seems to me that a great 
many people didn't vote for Trump because of who he was, but *in spite* 
of who he was - given the choice between two people who they perceived 
as corrupt, malevolent billionaires, they went for the one who wasn't 
part of the political machine they see as irrevocably broken, and at 
least said some things to indicate that he noticed them. Essentially a 
protest vote, just on a country-wide scale. I just hope the consequences 
won't be as severe as we all fear.


My personal answer to your final question comes in two parts:

(1) As I understand it, there's at least some indication that it isn't 
as bad as it seems (or, more precisely, it's always been at least this 
bad, but we just couldn't see it as clearly before). History is littered 
with extreme examples of anti-science gaining control at the highest 
levels of government (Lysenkoism, witch hunts, creationism, etc.). At 
least these days it seems science is mostly on the front foot 
(anti-science movements and politicians seem to be mostly fighting a 
slowly losing battle against concepts introduced by scientists, rather 
than scientists mostly fighting back against unscientific dogma).


(2) A big part of the solution has to be in communication. Our ability 
to communicate our work (and its importance) directly to the public has 
never been greater. That "directly" bit is important, because science 
journalists and even university PR departments have historically had a 
habit of both mangling the science and overblowing its impact between 
the scientist and the newsprint. Providing clear, accessible and, above 
all, friendly explanations of key work online can count for a heck of a 
lot.


Yes, we're certainly facing big problems - but I'm not yet ready to 
despair.


Best regards,

Tristan





On 2016-11-10 09:47, Elton Zeqiraj wrote:

Hi Tristan,

I’m afraid I don’t get the logic in the article you sent. In this
case the farmers who are crying for the attention of the people in
shiny palaces have voted for one who lives in golden mansions.

For me the really worrying thing here is how can so many people be
misled by a false prophet who lies and misbehaves so much. It is a
systemic problem in our society. We never before had so much access to
information, and yet it is so easy to mislead people.

People understood what Trump was and they basically said: “I don’t
care”! We are living in a world where people don’t care about
facts, they just say “I know it to be true”. As scientists whose
job and mission in life is to further and pass knowledge, it is very
serious that we are in this post-truth era. It affects us all when we
talk about climate change, evidence based treatments in our hospitals,
GM debate, etc.

Instead of making this about Trump, I would like to pose a different
question: How are we going to deal with the anti-expert movement that
is now so prominent in our society?

Cheers,
Elton


On Nov 10, 2016, at 8:15 AM, Tristan Croll <ti...@cam.ac.uk> wrote:

In the interests of promoting understanding... the link below is to
an article on what is ostensibly a comedy website and contains a bit
of coarse language, but nevertheless is quite possibly the most
insightful exposition of the situation I've come across. The
two-sentence synopsis: don't think of this as the forces of hate,
fear and ignorance winning. Think of it as a cry for attention from
a large number of people who are seriously struggling and (mostly
correctly) see their problems as being ignored by the system.
Writing them off as ignorant and hateful isn't the answer.

In agreement with various others, this is my first and last post
referencing politics or religion on this forum.



http://www.cracked.com/blog/6-reasons-trumps-rise-that-no-one-talks-about/

[1]

On 2016-11-10 08:00, Marjolein Thunnissen wrote:
Dear Bill,
I fully agree with you, awareness has to be spread and one should
not
ignore politics completely, especially when there are so strong
anti-intellectual (anti-science) statements out there.
best regards
Marjolein
On 10 Nov 2016, at 04:17, William G. Scott <wgsc...@ucsc.edu> wrote:
Dear Edward et al:
I agree we shouldn’t engage in partisan arguments on the CCP4bb.
However, I think it is a mistake, and perhaps a missed opportunity,
to ignore politics completely.
For example, Newt Gingrich is currently in the running for Sec HHS.
He has previously written editorials in the NYT and Wall Street
Journal advocating doubling the budget of the NIH.
I think it is incumbent upon us to make our voices heard if such an
opportun

Re: [ccp4bb] ligand binding and crystal form

2016-10-26 Thread Tristan Croll

Are these three crystals in order of harvesting (with different soaking times)? 
How big is your ligand. How accessible is the binding pocket (and is there a 
clear difference in accessibility between chains)?

T

 
 
Tristan Croll
Research Fellow
Cambridge Institute for Medical Research
University of Cambridge CB2 0XY
 

 

> On 26 Oct 2016, at 13:32, Veronica Fiorentino <veronicapfiorent...@gmail.com> 
> wrote:
> 
> Hello all,
> I just solved a NCS-tetrameric (biological assembly is just a dimer) crystal 
> structures with ligand soak (same plate - same conditions). No density for 
> ligand is observed in the first map. In the 2nd, I have 1 ligand bound. In 
> the 3rd, I have 2 ligands bound. Is there any reason for this 'random' 
> behaviour? 
> 
> In addition, I observed just one crystal out of 20 gave a different unit 
> cell. Pointless confirms to me 
> "Best Solution:space group C 2 2 2". REFMAC refinement shows R/Rfree ~ 
> 20/25 %
> Cell from mtz :   216.5   345.8   145.290.090.090.0
> Space group from mtz: number -   21; name - C 2 2 2
> 
> All other datasets have:
> Cell from mtz :   147.0   354.3   217.490.090.090.0
> Space group from mtz: number -   20; name - C 2 2 21
> 
> I tried re-processing/refining the C2221 dataset in C222 but R/Rfree stays 
> ~45%. Can I also consider the C2221 dataset as a 'different crystal form'?
> 
> Am I safe?
> 
> Thank you all,
> Veronica

Re: [ccp4bb] VR – I can’t believe no-one is talking about this!

2015-11-25 Thread Tristan Croll

There are a few visualisation packages out there that either have VR support 
now or are very close to having it in place. I've personally had a chance to 
briefly try out the Oculus Rift (aka seasickness simulator) in VMD. The 
challenge with these things is doing the head-tracking, re-calculating the 
display, and showing it fast enough that the delay doesn't cause 
disorientation/dizziness - or, alternatively, predicting head movements well 
enough so that the eyes see what they expect. It's getting very close, but 
still imperfect.

If you want to get really excited, though, check out the Microsoft Hololens and 
start imagining the possibilities.

Cheers,

Tristan

-Original Message-
From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Robert 
Sweet
Sent: Thursday, 26 November 2015 11:28 AM
To: CCP4BB@JISCMAIL.AC.UK
Subject: [ccp4bb] VR – I can’t believe no-one is talking about this!

Iʼm a synchrotron geek and Iʼve not actually solved a structure for years, and 
I certainly never ran COOT. The NY Times recently send all of us subscribers a 
stereo viewer: http://www.google.com/get/cardboard/  It came with our Sunday 
paper.  It helped introduce a new NYT feature, which is virtual-reality news 
stories.  Here is the website presenting this feature as it is today: 
http://www.nytimes.com/newsgraphics/2015/nytvr/   The initial feature was the 
one entitled The Displaced.

I donʼt happen to own a Smart Phone, but my son Charley (a photographer) does 
and probably most of you do. He hadnʼt really learned about this, but he 
happens to be home for Thanksgiving and tried it out. I watched the full 
episode entitled Backwater on the nytvr web site. It's pretty thrilling!

To me the idea of touring the inside of a protein molecule pops up instantly.  
Back in the early days of stereo images for computerized model building, folks 
would talk about "virtual reality."  To me this wasn't virtual, it was REALITY; 
I WAS inside the molecule looking around.  Is any of you looking at this 
smartphone/stereo viewer combo?  Why is there no chatter on ccp4bb?  What a 
great way to recruit grad students!

Bob

=
    Robert M. Sweet   E-Dress: sw...@bnl.gov
    Principal Investigator, LSBR: The Life Science ^ (that's L
      and Structural Biology Resource at NSLS-II   not 1)
    Photon Sciences and Biosciences Dept
    Office and mail, Bldg 745, a.k.a. LOB-5
    Brookhaven Nat'l Lab. Phones:
    Upton, NY  11973  631 344 3401  (Office)
    U.S.A.    631 344 2741  (Facsimile)

=

Re: [ccp4bb] Classifying Diverse Conformations of Many Structures of a Protein

2015-07-07 Thread Tristan Croll

Sounds like a perfect application for principal components analysis. Check out 
Bio3D: http://thegrantlab.org/bio3d/index.php.

Cheers,

Tristan

 
 
Tristan Croll
Lecturer
Faculty of Health
School of Biomedical Sciences
Institute of Health and Biomedical Engineering
Queensland University of Technology
60 Musk Ave
Kelvin Grove QLD 4059 Australia
+61 7 3138 6443
 
This email and its attachments (if any) contain confidential information 
intended for use by the addressee and may be privileged.  We do not waive any 
confidentiality, privilege or copyright associated with the email or the 
attachments.  If you are not the intended addressee, you must not use, 
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 On 8 Jul 2015, at 11:06 am, Keller, Jacob kell...@janelia.hhmi.org wrote:
 
 Is anyone aware of a way to classify large numbers (100s) of 
 conformationally-diverse crystal structures of a single protein (here 
 calmodulin)? Pairwise RMSD matrixes seem possible, but may be complicated 
 since there are two somewhat stable lobes, and the flexible linker in the 
 middle. What I am imagining is a sort of multidimensional tree depicting the 
 relationships in conformation space of the various structures.
 
 I remember something for this called esct or similar, but can't seem to 
 google it.
 
 Any thoughts?
 
 Jacob
 
 ***
 Jacob Pearson Keller, PhD
 Looger Lab/HHMI Janelia Research Campus
 19700 Helix Dr, Ashburn, VA 20147
 email: kell...@janelia.hhmi.org
 ***

Re: [ccp4bb] Cis-peptide bond checking

2015-07-03 Thread Tristan Croll


Hi all,

Just revisiting this old thread, since it would seem I owe Paul Emsley 
something of an apology... after discussion with various people prior to 
submission of my manuscript, I concluded that overly-aggressive manual model 
building in Coot was probably the most common source of erroneous cis bonds, 
and said as much in the text. Now I'm thinking they probably creep in much 
earlier. I've just been looking at the output of an AutoBuild run of a 3.1A 
structure (my first ever - I'm not a crystallographer by training, so I'm sort 
of working backwards from the improvement of existing structures to learning 
how to produce them), and I count 16 cis bonds in 640 residues. If I'm reading 
the code correctly, I think the problem arises in RESOLVE (specifically, in 
assemble_model.cpp) when individual fragments are joined - overlapping 
fragments are trimmed back and ligated, but there's never a geometric check for 
the stereochemistry of the newly formed peptide bond. So, any time the data 
becomes a bit ambiguous there's a chance that one of the two residues at the 
join will be in a sufficiently incorrect orientation that the peptide bond is 
made the wrong way around.

Best regards,

Tristan


From: Tristan Croll
Sent: Tuesday, 17 February 2015 6:13 AM
To: wouter.t...@radboudumc.nl; CCP4 bulletin board
Subject: Re: [ccp4bb] Cis-peptide bond checking


Dear Wouter,


That does sound like a useful tool indeed - finding the proverbial needle in a 
haystack! That's the challenge with such a rare event: rather like a true 
Ramachandran outlier, when they do occur they're usually a sign of an important 
motif in your protein that should be remarked upon.


To others making the same point: yes, I'm well aware of the existence of true 
cis peptides, and both re-calculate the background rate in high-res structures 
and briefly discuss their nature in my paper (my personal favourite example is 
tissue transglutaminase (2q3z) which contains two - one of which is induced by 
the formation of a vicinal disulfide bond. It's believed that reduction of the 
disulfide switches the backbone back to trans to activate the enzyme). But I'm 
currently unaware of any protein that contains more than 3-4 cis bonds that 
stand up under scrutiny, while there are many models out there with tens of, or 
up to a few hundred.  For examples of erroneous assignment at high res look at 
3ncq, 2gec or 2j82.


It's not such a problem at high resolution, but at lower resolutions I'm more 
concerned about why the cis bonds have crept into the model. Are they simple 
innocuous oversights (as pointed out by Robbie Joosten, most - but certainly 
not all - appear in poorly-defined density), or have they come about due to 
accidentally force-fitting a loop that is fundamentally wrong (e.g. due to an 
adjacent strand being out of register)? In most cases it's of course the 
former, but what worries me is the example of a structure I found (since 
corrected by the authors) that had 86 cis bonds (1.4%), yet only 0.4% 
Ramachandran and RSRZ outliers. In a good structure one would expect an 
erroneous cis bond to introduce an outlier in some other metric - but it seems 
equally possible that in a bad structure it could bring an outlier back into 
a favoured region.


Hope this clarifies my point.


Cheers,


Tristan


From: wouter.t...@radboudumc.nl wouter.t...@radboudumc.nl
Sent: Monday, 16 February 2015 9:55 PM
To: Tristan Croll; CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] Cis-peptide bond checking

Dear Tristan,

Thank you for your post of earlier today regarding the problem of cis and trans 
peptide planes in the PDB. We also realised this problem a while ago and an 
article describing this problem and a solution is presently under review at 
Acta Cryst. D. After analysis of the PDB we can state with 95% certainty that 
~4600 trans - cis flips in ~2800 entries (and ~70K peptide-plane flips) are 
needed in the PDB. Around a third of the trans - cis corrections concern 
non-prolines. We hope to be able to deal with the problem of cis - trans 
corrections later.
In the tradition of our group, the software to detect these flips is already 
available at swift.cmbi.ru.nl.

Hopefully, the referees of our article consider this topic just as important as 
you and I do :-).

Kind regards,

Wouter Touw and Gert Vriend


On 02/16/2015 10:58 AM, Tristan Croll wrote:

Dear all,


My apologies for the spam-like nature of my post, but I would like to draw your 
attention to an important issue (outlined in an upcoming short communication to 
Acta D, which will appear at doi:10.1107/S1399004715000826 once it's online). 
At present, neither the structural quality checks in commonly-used 
crystallography packages nor those run on deposition of a structure to the PDB 
are flagging the presence of non-proline cis peptide bonds. This has led to the 
presence of many erroneous cis bonds creeping

Re: [ccp4bb] Tris buffer in cryo protectant

2015-06-12 Thread Tristan Croll

Might be worth trying to see if your protein will still crystallize in a 
mixture of tris and TAPS buffer? The pKa of the latter is very close to tris, 
but goes in the opposite direction with temperature - a roughly 3:2 TAPS:tris 
mix should have minimal pH change on freezing.

 
 
Tristan Croll
Lecturer
Faculty of Health
School of Biomedical Sciences
Institute of Health and Biomedical Engineering
Queensland University of Technology
60 Musk Ave
Kelvin Grove QLD 4059 Australia
+61 7 3138 6443
 
This email and its attachments (if any) contain confidential information 
intended for use by the addressee and may be privileged.  We do not waive any 
confidentiality, privilege or copyright associated with the email or the 
attachments.  If you are not the intended addressee, you must not use, 
transmit, disclose or copy the email or any attachments.  If you receive this 
email by mistake, please notify the sender immediately and delete the original 
email.
 
 

 On 13 Jun 2015, at 6:49 am, Ursula Schulze-Gahmen uschulze-gah...@lbl.gov 
 wrote:
 
 Does anyone have experience with Tris buffer in cryo protectants? I would 
 expect the pH of the cryosolution to increase a lot during flash freezing 
 which could perhaps destroy the diffraction. I rarely use Tris for 
 crystallization but the current protein really prefers Tris. I would 
 appreciate any comments.
 
 Ursula
 
 -- 
 Ursula Schulze-Gahmen, Ph.D.
 Project Scientist
 UC Berkeley, QB3
 360 Stanley Hall #3220
 Berkeley, CA 94720-3220
 (510) 643 9491

Re: [ccp4bb] Should I refine a low-resolution dataset 6 A

2015-06-09 Thread Tristan Croll

This is a very tough resolution to be working at, but if you want to take it on 
I'd suggest that xMDFF 
(http://www.ks.uiuc.edu/Training/Tutorials/science/mdff/tutorial_mdff-html/node8.html)
 starting from a good homology model is more likely to yield results. Shameless 
self-promotion: I've been working with the xMDFF people (particularly Ryan) to 
allow interactive MD-based remodeling of key regions using a haptic interface, 
which is proving very powerful (and fun!) in the 3-4 Angstrom range. The first 
paper using this is submitted, and the software should be available with the 
next release of VMD... stay tuned!

At 6 Angstroms, it's hard to say how much you'll get. If your starting homology 
model is good and your data reasonably complete, you may not do too badly - but 
ultimately you're going to be far more dependent on judgement based on factors 
other than the density (i.e. interatomic forces) than you would be at higher 
resolution. At a resolution like this I'd be running my model in implicit 
solvent, with secondary structure restraints applied and only weak coupling to 
the map - basically everything geared to prevent trapping in local minima.

Hope this helps,

Tristan



Tristan Croll
Lecturer
Faculty of Health
School of Biomedical Sciences
Institute of Health and Biomedical Engineering
Queensland University of Technology
60 Musk Ave
Kelvin Grove QLD 4059 Australia
+61 7 3138 6443

This email and its attachments (if any) contain confidential information 
intended for use by the addressee and may be privileged.  We do not waive any 
confidentiality, privilege or copyright associated with the email or the 
attachments.  If you are not the intended addressee, you must not use, 
transmit, disclose or copy the email or any attachments.  If you receive this 
email by mistake, please notify the sender immediately and delete the original 
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On 9 Jun 2015, at 9:31 pm, Kerstin Keller 
kerstin.keller.1...@outlook.commailto:kerstin.keller.1...@outlook.com wrote:

Hi Experts,

Last year I collected a dataset at 6 A of 2600 aa multi-domain protein.

Resolution range 50 - 6 (6.7-6)
Wilson B370 A^2
Reflection 34120 (12240)
Rmeas 0.2 (4.4)
Rpim 0.06(1.2)
I-sigI  9 (1)
CC1/2   0.9 (0.5)

I have the following questions in my mind.

1. Does it make any sense to solve the structure at _this_ resolution? It is 
not completely novel protein, there are known structures with about 54 % 
identity to this. The fold is known to be the same.

2. Doing a molecular replacement with Phaser using EM-model gives me a unique 
solution. And I can see a reasonable electron density map. Tried again with 
AMORE - The amore-build-output model also gives the same solution. In both 
cases, the solutions are unique. There are no so-called translational symmetry.

3. If I do a Refmac restrained refinement, though I get R/Rfree in around 
30.1/35.5 the stereochemistry is very poor (18 % outliers). I had to enable 
tight WEIGHT MATRIX (1e-7). Here at this resolution does it make any sense to a 
restrained refinement?

4. If I do only a rigid body with Refmac, the R-factor/Rfree are at around 41 
%, and in many places model does not fit density. When I manually correct these 
and refine there is basically no change in R-factor/R-free (it even worsens in 
cases).

From: Bernhard Rupp (Hofkristallrat a.D.) 
hofkristall...@gmail.commailto:hofkristall...@gmail.comDate: 27 April 2015 
at 21:54
Subject: Re: [ccp4bb] 3BDN, 16.5% Ramachandran Outliers!
To: CCP4BB@jiscmail.ac.ukmailto:CCP4BB@jiscmail.ac.uk
What we cannot tell sans supporting density is whether it is a more accurate 
model, although I haverarely seen an improvement in geometry giving worse 
density fit. Usually a mess remains a mess -
there is (at this resolution) no free lunch. The key question is again – does 
the model justify

5. I did try DEN, reference model restraints, PROSMART(refmac) etc which have 
no improvement at all - R/Rfree stuck at 41 %.

6. Since my group is EM-group, I wonder when EM-maps of 6 A are published, why 
are X-ray data at the same resolution not being published? What happens to 
these datasets?

7. Can I _just_ do a molecular replacement and just mutate residues (based on a 
sequence alignment - There are large numbers of deletions and hence sequence 
registers are different/unknown) and deposit it as a model in the PDB? Should I 
put the side-chains or it is meaning less at this resolution? Why in the 
EM-field they are allowed to deposit such coordinates with side-chains?

8. As hofkristall...@gmail.commailto:hofkristall...@gmail.com points out

Particularly in Molecular Replacement structures, and here particularly in 
those with multi-segment/domain models, there are almost always parts that fit 
well and othersthat fit poorly -  with simply not enough data at the given 
resolution to improve the poor partssans additional phase information. Bias 
issues have been discussed and need not be iterated here.
Since my protein also has

Re: [ccp4bb] modified amino acids

2015-05-14 Thread Tristan Croll

A chemical component search through the Chemical Component Dictionary 
(http://www.rcsb.org/pdb/ligand/chemAdvSearch.do) should get you there fairly 
quickly. Just draw in the core amino acid N-CA(CB)-C=O and you should end up 
with a pretty comprehensive list (although you'll need to handle glycine 
separately). You'll probably end up needing to prune out a lot of non-amino 
acids, but that shouldn't be horribly arduous. An alternative approach might be 
to search by name at HIC-Up 
http://xray.bmc.uu.se/hicup/.http://xray.bmc.uu.se/hicup/


Cheers,


Tristan



From: CCP4 bulletin board CCP4BB@JISCMAIL.AC.UK on behalf of Debasish 
Chattopadhyay debas...@uab.edu
Sent: Friday, 15 May 2015 9:46 AM
To: CCP4BB@JISCMAIL.AC.UK
Subject: [ccp4bb] modified amino acids


I am looking for a list of all modified amino acids found in protein structures 
(in PDB).  Three letter codes and geometry files will be wonderful.



Thanks



Debasish

Re: [ccp4bb] cryo condition

2015-05-04 Thread Tristan Croll

What about nature's favourite cryoprotectant, trehalose?

From: CCP4 bulletin board CCP4BB@JISCMAIL.AC.UK on behalf of Roger Rowlett 
rrowl...@colgate.edu
Sent: Tuesday, 5 May 2015 7:22 AM
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] cryo condition

We rarely use glycerol anymore, because it seems to fail so often for
many of our current proteins. Try glucose, 25-30%. This is most
conveniently done by weighing 125-150 mg of glucose in a microcentrifuge
tube, then addding well solution to the 0.5 mL mark and mixing until
completely dissolved. Then you can try dunking crystals in the cryo
solution, or, you can try the no-fail method (which does fail on
occasion) of cryoprotecting directly in the crystallization drop by slow
addition of the cryoprotectant. See
http://capsicum.colgate.edu/chwiki/tiki-index.php?page=Mounting+Protein+Crystals.
We have often found the slow addition of a glucose cryoprotectant works
for fragile, high solvent content crystals that are prone to cracking
under osmotic stress.

Other alternatives include high concentrations of sodium malonate
(1-2M), or high concentrations of sodium formate (I think around 4 M?).
These could also be introduced gradually if required.

Good luck!

___
Roger S. Rowlett
Gordon  Dorothy Kline Professor
Department of Chemistry
Colgate University
13 Oak Drive
Hamilton, NY 13346

tel: (315)-228-7245
ofc: (315)-228-7395
fax: (315)-228-7935
email: rrowl...@colgate.edu

On 5/4/2015 2:43 PM, Faisal Tarique wrote:
 Hello everyone

 Can anybody suggest me a cryo condition for a crystal obtained in
 MIDAS screen of Molecular Dimension:

 G1 0.1MTris8.0G10.1Mpotassium chloride25% v/vSOKALAN®CP7 0.1MHEPES7.0

 G20.3Mammonium formate20% v/vSOKALAN® CP 5 0.1MHEPES7.0

 Crystals are in beautiful cuboid shaped but all sorts of PEG
 combinations and Glycerol formulation failed to prevent it from
 cracking and dissolving.

 Has anybody faced a similar situation as mentioned above and what
 precaution was taken to prevent it from cracking or dissolving.

 Your suggestions will be of immense help

 Thanks in advance

Re: [ccp4bb] Off-topic - Crystallisation in anaerobic glove box

2015-03-18 Thread Tristan Croll

It's a little complicated. It's true that oxygen is more soluble in most oils 
than in water - but in a high viscosity mineral oil the diffusion rate is 
orders of magnitude lower. So the combination of an oil overlay and a reducing 
agent in your buffer should protect your sample much longer than the reducing 
agent alone - as long as your oil was degassed to start with. Note that silicon 
oils are a bad choice for this - silicones have an enormous affinity for oxygen 
(so much so that they've been explored as artificial blood substitutes), and it 
diffuses through them very readily.

 
 
Tristan Croll
Lecturer
Faculty of Health
School of Biomedical Sciences
Institute of Health and Biomedical Engineering
Queensland University of Technology
60 Musk Ave
Kelvin Grove QLD 4059 Australia
+61 7 3138 6443
 
This email and its attachments (if any) contain confidential information 
intended for use by the addressee and may be privileged.  We do not waive any 
confidentiality, privilege or copyright associated with the email or the 
attachments.  If you are not the intended addressee, you must not use, 
transmit, disclose or copy the email or any attachments.  If you receive this 
email by mistake, please notify the sender immediately and delete the original 
email.
 
 

 On 18 Mar 2015, at 11:49 pm, Edward A. Berry ber...@upstate.edu wrote:
 
 Do you have evidence that the oil blocks diffusion of O2? O2 is a nonpolar 
 molecule, generally much more soluble in oils than in water. I'm not sure 
 about silicone oils, but I would think they also dissolve O2 readily.
 eab
 
 On 03/18/2015 08:02 AM, Patrick Shaw Stewart wrote:
 
 Hi Steve
 
 I have one more comment for this thread.
 
 The microbatch-under-oil method is very handy for anaerobic work:
 
1.  You can keep the microbatch stock solutions in normal microtitre 
 plates (polypropylene is best to reduce evaporation) for months, which 
 hugely reduces the amount of degassing that you need to do.  You will only 
 use say 0.5 ul of stock per drop.
 
2.  The oil offers a surprising amount of protection from oxidation, 
 which may be helpful eg in harvesting.
 
3.  Microbatch can be automated - in parallel to vapor diffusion if 
 desired
 
 
 It's amazing how often (aerobic) microbatch produces far superior crystals 
 to V.D. for no obvious reason - it's well worth trying for both screening 
 and optimization.
 
 Best wishes
 
 Patrick
 
 
 
 On 11 March 2015 at 10:17, Stephen Carr stephen.c...@rc-harwell.ac.uk 
 mailto:stephen.c...@rc-harwell.ac.uk wrote:
 
Dear CCP4BBer's
 
Apologies for the off-topic post, but the CCP4BB seems to be the best 
 place to ask about crystallisation.
 
I am looking to set up crystallisation in an anaerobic glove box and 
 wondered how other people did this, specifically the crystallisation stage.  
 My initial thoughts were to place a small crystallisation incubator inside 
 the box, however the smallest I have come across so far (~27L) is still 
 rather large.  Has anyone come across smaller incubators?  Alternatively are 
 incubators even neccessary if the glove box is placed in a room with good 
 air conditioning and stable temperature control?
 
Any recommendations would be very helpful.
 
Thanks in advance,
 
Steve Carr
 
Dr Stephen Carr
Research Complex at Harwell (RCaH)
Rutherford Appleton Laboratory
Harwell Oxford
Didcot
Oxon OX11 0FA
United Kingdom
Email stephen.c...@rc-harwell.ac.uk mailto:stephen.c...@rc-harwell.ac.uk
tel 01235 567717 tel:01235%20567717
 
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Re: [ccp4bb] Cis-peptide bond checking

2015-02-16 Thread Tristan Croll

Dear Wouter,


That does sound like a useful tool indeed - finding the proverbial needle in a 
haystack! That's the challenge with such a rare event: rather like a true 
Ramachandran outlier, when they do occur they're usually a sign of an important 
motif in your protein that should be remarked upon.


To others making the same point: yes, I'm well aware of the existence of true 
cis peptides, and both re-calculate the background rate in high-res structures 
and briefly discuss their nature in my paper (my personal favourite example is 
tissue transglutaminase (2q3z) which contains two - one of which is induced by 
the formation of a vicinal disulfide bond. It's believed that reduction of the 
disulfide switches the backbone back to trans to activate the enzyme). But I'm 
currently unaware of any protein that contains more than 3-4 cis bonds that 
stand up under scrutiny, while there are many models out there with tens of, or 
up to a few hundred.  For examples of erroneous assignment at high res look at 
3ncq, 2gec or 2j82.


It's not such a problem at high resolution, but at lower resolutions I'm more 
concerned about why the cis bonds have crept into the model. Are they simple 
innocuous oversights (as pointed out by Robbie Joosten, most - but certainly 
not all - appear in poorly-defined density), or have they come about due to 
accidentally force-fitting a loop that is fundamentally wrong (e.g. due to an 
adjacent strand being out of register)? In most cases it's of course the 
former, but what worries me is the example of a structure I found (since 
corrected by the authors) that had 86 cis bonds (1.4%), yet only 0.4% 
Ramachandran and RSRZ outliers. In a good structure one would expect an 
erroneous cis bond to introduce an outlier in some other metric - but it seems 
equally possible that in a bad structure it could bring an outlier back into 
a favoured region.


Hope this clarifies my point.


Cheers,


Tristan


From: wouter.t...@radboudumc.nl wouter.t...@radboudumc.nl
Sent: Monday, 16 February 2015 9:55 PM
To: Tristan Croll; CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] Cis-peptide bond checking

Dear Tristan,

Thank you for your post of earlier today regarding the problem of cis and trans 
peptide planes in the PDB. We also realised this problem a while ago and an 
article describing this problem and a solution is presently under review at 
Acta Cryst. D. After analysis of the PDB we can state with 95% certainty that 
~4600 trans - cis flips in ~2800 entries (and ~70K peptide-plane flips) are 
needed in the PDB. Around a third of the trans - cis corrections concern 
non-prolines. We hope to be able to deal with the problem of cis - trans 
corrections later.
In the tradition of our group, the software to detect these flips is already 
available at swift.cmbi.ru.nl.

Hopefully, the referees of our article consider this topic just as important as 
you and I do :-).

Kind regards,

Wouter Touw and Gert Vriend


On 02/16/2015 10:58 AM, Tristan Croll wrote:

Dear all,


My apologies for the spam-like nature of my post, but I would like to draw your 
attention to an important issue (outlined in an upcoming short communication to 
Acta D, which will appear at doi:10.1107/S1399004715000826 once it's online). 
At present, neither the structural quality checks in commonly-used 
crystallography packages nor those run on deposition of a structure to the PDB 
are flagging the presence of non-proline cis peptide bonds. This has led to the 
presence of many erroneous cis bonds creeping into the PDB - primarily in 
low-resolution structures as one would expect, but I have identified clearly 
erroneous examples in structures with resolutions as high as 1.3 Angstroms. 
From my analysis, I estimate that a few thousand structures have been affected 
to some extent, with the worst cases having as high as 3% of their peptide 
bonds in cis. Particularly if you have published anything 2.5 Angstroms in the 
past few years, may I gently suggest that you make a quick double-check of your 
deposited structures? This can be done quickly and simply in Coot 
(Extensions-Modelling-Residues with Cis peptide bonds).


Best regards,


Tristan



Het Radboudumc staat geregistreerd bij de Kamer van Koophandel in het 
handelsregister onder nummer 41055629.
The Radboud university medical center is listed in the Commercial Register of 
the Chamber of Commerce under file number 41055629.

[ccp4bb] Cis-peptide bond checking

2015-02-16 Thread Tristan Croll

Dear all,


My apologies for the spam-like nature of my post, but I would like to draw your 
attention to an important issue (outlined in an upcoming short communication to 
Acta D, which will appear at doi:10.1107/S1399004715000826 once it's online). 
At present, neither the structural quality checks in commonly-used 
crystallography packages nor those run on deposition of a structure to the PDB 
are flagging the presence of non-proline cis peptide bonds. This has led to the 
presence of many erroneous cis bonds creeping into the PDB - primarily in 
low-resolution structures as one would expect, but I have identified clearly 
erroneous examples in structures with resolutions as high as 1.3 Angstroms. 
From my analysis, I estimate that a few thousand structures have been affected 
to some extent, with the worst cases having as high as 3% of their peptide 
bonds in cis. Particularly if you have published anything 2.5 Angstroms in the 
past few years, may I gently suggest that you make a quick double-check of your 
deposited structures? This can be done quickly and simply in Coot 
(Extensions-Modelling-Residues with Cis peptide bonds).


Best regards,


Tristan

Re: [ccp4bb] Cis-peptide bond checking

2015-02-16 Thread Tristan Croll

Yes, but when the validation reports are as comprehensive as 
thishttp://www.wwpdb.org/validation/validation-reports.php, it's all too easy 
to assume that they've covered all possible issues and that the lack of a list 
of cis bonds means there are none. I understand that the next release of 
MolProbity will include this in its suite of checks so the issue should begin 
to go away as people upgrade.


Cheers,


Tristan


From: Bernhard Rupp hofkristall...@gmail.com on behalf of Bernhard Rupp 
b...@ruppweb.org
Sent: Monday, 16 February 2015 8:55 PM
To: Tristan Croll; CCP4BB@JISCMAIL.AC.UK
Subject: RE: [ccp4bb] Cis-peptide bond checking

HmmCISPEP records are provided...

CISPEP   1 GLY A   63LEU A   64  0 1.59
CISPEP   2 LEU A   69PRO A   70  0   -14.05
CISPEP   3 PRO A   70PRO A   71  0-2.41
CISPEP   4 ASP A  162GLY A  163  0-8.79

Which can be parsed/checked
but then again, who does?

https://www.youtube.com/watch?v=bAZqE2DnxUI

Best, BR

From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Tristan 
Croll
Sent: Montag, 16. Februar 2015 10:58
To: CCP4BB@JISCMAIL.AC.UK
Subject: [ccp4bb] Cis-peptide bond checking


Dear all,



My apologies for the spam-like nature of my post, but I would like to draw your 
attention to an important issue (outlined in an upcoming short communication to 
Acta D, which will appear at doi:10.1107/S1399004715000826 once it's online). 
At present, neither the structural quality checks in commonly-used 
crystallography packages nor those run on deposition of a structure to the PDB 
are flagging the presence of non-proline cis peptide bonds. This has led to the 
presence of many erroneous cis bonds creeping into the PDB - primarily in 
low-resolution structures as one would expect, but I have identified clearly 
erroneous examples in structures with resolutions as high as 1.3 Angstroms. 
From my analysis, I estimate that a few thousand structures have been affected 
to some extent, with the worst cases having as high as 3% of their peptide 
bonds in cis. Particularly if you have published anything 2.5 Angstroms in the 
past few years, may I gently suggest that you make a quick double-check of your 
deposited structures? This can be done quickly and simply in Coot 
(Extensions-Modelling-Residues with Cis peptide bonds).



Best regards,


Tristan

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