Re: [ccp4bb] Fw:[ccp4bb] on Cell & Symmetry in coot

[Martyn Symmons]

that's neat - I have used chimera to build filaments from fibre
diffraction where you specify the rise and angle rather than symmops.
'Sym' command specifying 'group h' then rise and angle.
Martyn

On Fri, Dec 9, 2016 at 1:34 PM, Paul Emsley <pems...@mrc-lmb.cam.ac.uk> wrote:
>
> I would use the function new_molecule_by_symop
>
> e.g.
>
> # python
> imol = new_molecule_by_symop(0, "Y,-X+Y,Z+1/6", 0,0,0)
>
> then do this 5 more times selecting the other symmetry operators that
> generate the helix around the z axis..
>
> or the attached script (adjust as necessary)
>
> Paul.
>
>
> On 09/12/16 13:06, Martyn Symmons wrote:
>>
>> Yes I would do it that way by clicking on the ones I want to see -
>> then saving them and then reading them back in. You can change the
>> chainids so Coot makes them different colours which might help.
>> Things might be easier in PyMol if you want interactive control over
>> the colours for a picture say.
>> cheers
>>   Martyn
>>
>> On Fri, Dec 9, 2016 at 12:59 PM, Smith Liu <smith_liu...@163.com> wrote:
>>>
>>> Dear All,
>>>
>>> I mean if the radius set in the Coot "Cell Symmetry" was too small, not
>>> enough monomers (less than 6) can be displayed to show the "continuous
>>> helix
>>> with a six-fold screw axis". If the radius was too large, as for the
>>> "continuous helix with a six-fold screw axis" can be regarded as a "rod",
>>> too large radius will lead to show several rods in one window. But with
>>> the
>>> Coot window, it cannot distinguish which monomer was from which rod. Thus
>>> I
>>> cannot identify 6 monomers forming the single rod, i.e., a  "continuous
>>> helix with a six-fold screw axis".
>>>
>>> Can anyone explain in this situation how can I identify the 6 monomers in
>>> the Coot "Cell and SYmmetry" windows forming a single "continuous helix
>>> with
>>> a six-fold screw axis"?
>>>
>>> Smith
>>>
>>>
>>>
>>>
>>>
>>>  Forwarding messages 
>>> From: "Smith Lee" <0459ef8548d5-dmarc-requ...@jiscmail.ac.uk>
>>> Date: 2016-12-09 18:12:20
>>> To: CCP4BB@JISCMAIL.AC.UK
>>> Subject: [ccp4bb] on Cell & Symmetry in coot
>>>Dear All,
>>>
>>> There is a pdb, once opended in coot, it was a monomer (space group   P
>>> 65 2
>>> 2 ). But in the correspondence paper, it writes, "the subunits form a
>>> continuous helix with a six-fold screw axis".
>>>
>>> I have tried to view with Coot the "six-fold screw axis" formed by 6
>>> monomers. But in the "Cell & Symmetry" in Coot, if the radius is small, 6
>>> monomers cannot be shown. If I increase the radius, more than 6 monomers
>>> would occur in the window, and it can hardly distinguish the 6 monomers
>>> forming the "six-fold screw axis".
>>>
>>> In this situation, will you please let me know how to use Coot to
>>> identify
>>> the 6 monomers forming the "six-fold screw axis"? In addition, suppose  6
>>> monomers forming the "six-fold screw axis" have been identified in Coot,
>>> in
>>> order to save the pdb of each monomer, I need to click each monomer in
>>> mouse, then by "Save symmetry coordinates" to save the pdb of each
>>> monomer,
>>> right?
>>>
>>> I am looking forward to getting your reply.
>>>
>>> Smith
>>>
>>>
>>>
>

Re: [ccp4bb] Fw:[ccp4bb] on Cell & Symmetry in coot

[Martyn Symmons]

Yes I would do it that way by clicking on the ones I want to see -
then saving them and then reading them back in. You can change the
chainids so Coot makes them different colours which might help.
Things might be easier in PyMol if you want interactive control over
the colours for a picture say.
cheers
 Martyn

On Fri, Dec 9, 2016 at 12:59 PM, Smith Liu  wrote:
> Dear All,
>
> I mean if the radius set in the Coot "Cell Symmetry" was too small, not
> enough monomers (less than 6) can be displayed to show the "continuous helix
> with a six-fold screw axis". If the radius was too large, as for the
> "continuous helix with a six-fold screw axis" can be regarded as a "rod",
> too large radius will lead to show several rods in one window. But with the
> Coot window, it cannot distinguish which monomer was from which rod. Thus I
> cannot identify 6 monomers forming the single rod, i.e., a  "continuous
> helix with a six-fold screw axis".
>
> Can anyone explain in this situation how can I identify the 6 monomers in
> the Coot "Cell and SYmmetry" windows forming a single "continuous helix with
> a six-fold screw axis"?
>
> Smith
>
>
>
>
>
>  Forwarding messages 
> From: "Smith Lee" <0459ef8548d5-dmarc-requ...@jiscmail.ac.uk>
> Date: 2016-12-09 18:12:20
> To: CCP4BB@JISCMAIL.AC.UK
> Subject: [ccp4bb] on Cell & Symmetry in coot
>   Dear All,
>
> There is a pdb, once opended in coot, it was a monomer (space group   P 65 2
> 2 ). But in the correspondence paper, it writes, "the subunits form a
> continuous helix with a six-fold screw axis".
>
> I have tried to view with Coot the "six-fold screw axis" formed by 6
> monomers. But in the "Cell & Symmetry" in Coot, if the radius is small, 6
> monomers cannot be shown. If I increase the radius, more than 6 monomers
> would occur in the window, and it can hardly distinguish the 6 monomers
> forming the "six-fold screw axis".
>
> In this situation, will you please let me know how to use Coot to identify
> the 6 monomers forming the "six-fold screw axis"? In addition, suppose  6
> monomers forming the "six-fold screw axis" have been identified in Coot, in
> order to save the pdb of each monomer, I need to click each monomer in
> mouse, then by "Save symmetry coordinates" to save the pdb of each monomer,
> right?
>
> I am looking forward to getting your reply.
>
> Smith
>
>
>

Re: [ccp4bb] on Cell & Symmetry in coot

[Martyn Symmons]

Under the coot Show symmetry option - switch it to 'Symmetry by
Molecule> Display as CA' - (click on the Symmetry by Molecule menu
item to bring up the Symmetry Controller options window).

Unlike the  'Display sphere' option, with 'Display as CA' you can
increase the Symmetry Atom Display Radius by a large number and get a
nice simple view of the packing even over several cells.

(there is also a colour by symmetry option in the Symmetry Controller window)
The clicking on a chain to save still works really well for the CA
display version of your packing (kudos to Paul Emsley for this tool).

all the best
 Martyn

Martyn Symmons
Cambridge


On Fri, Dec 9, 2016 at 10:12 AM, Smith Lee
<0459ef8548d5-dmarc-requ...@jiscmail.ac.uk> wrote:
>   Dear All,
>
> There is a pdb, once opended in coot, it was a monomer (space group   P 65 2
> 2 ). But in the correspondence paper, it writes, "the subunits form a
> continuous helix with a six-fold screw axis".
>
> I have tried to view with Coot the "six-fold screw axis" formed by 6
> monomers. But in the "Cell & Symmetry" in Coot, if the radius is small, 6
> monomers cannot be shown. If I increase the radius, more than 6 monomers
> would occur in the window, and it can hardly distinguish the 6 monomers
> forming the "six-fold screw axis".
>
> In this situation, will you please let me know how to use Coot to identify
> the 6 monomers forming the "six-fold screw axis"? In addition, suppose  6
> monomers forming the "six-fold screw axis" have been identified in Coot, in
> order to save the pdb of each monomer, I need to click each monomer in
> mouse, then by "Save symmetry coordinates" to save the pdb of each monomer,
> right?
>
> I am looking forward to getting your reply.
>
> Smith

Re: [ccp4bb] Superpose program in CCP4

[Martyn Symmons]

I think superpose does not output CAs outwith a certain cut-off based
on the quality of the superimposition. It does try to match as many as
give reasonable RMSDs - but it is mainly focussed on the residues in
matching secondary elements as this is where it starts the
superimposition.

A simpler approach is to use LSQKAB which will report all CA RMSDs in
a file if you specify that you want DELTAS. But notice that as Gert
said this will probably give you a different kind of superimposition.
A workaround would be to restrict LSQKAB to the residue ranges
SUPERPOSE used - then it will produce the same superimposition but can
report DELTAS for _all_ residues (! phew).

A non-CCP4 approach is Chimera Multi-align Viewer which calculates
RMSD without doing any superimposition. So you could run it on your
SUPERPOSE output coordinate files. I recall RMSD is a 'header' in that
tool and can be output to a file.
https://www.cgl.ucsf.edu/chimera/docs/ContributedSoftware/multalignviewer/framemav.html
(You have to read your structures into Chimera before launching
Multi-align viewer).

hope that helps.
 Martyn

Martyn Symmons
Cambridge

On Sun, Oct 30, 2016 at 10:54 AM, Anastassis Perrakis <a.perra...@nki.nl> wrote:
> Dear Wenhe,
>
> Besides this advice, have a look at the
> http://webapps.embl-hamburg.de/rapido/ server.
>
> Sometimes its goodo to re-think of what you want to do, and wonder why its
> not easily doable in software (perhaps because its not the right thing to do
> …)
>
> A.
>
>
> On 30 Oct 2016, at 11:45, Gert Vriend <gerrit.vri...@radboudumc.nl> wrote:
>
> Dear Wenhe,
>
> No 3D superpose tool will always align/map all Calphas. If in the one
> protein the loop turns left, and in the other it turns right, then mapping
> those loops is meaningless and thus not done by good software. The other
> problem is that often two proteins that get compared do not even have
> equally many residues so that there will always be some unaligned/unmapped
> Calphas left at the end. Look for some articles by Arthur M Lesk on this
> topic, he has explained protein superposition (problems) very clearly.
>
> Gert
>
> Ps, if you want proteins superposed and get different output from what the
> standard software gives you, just mail me those PDB files and I can see what
> I can do.
>
>
> On 29-10-2016 17:47, WENHE ZHONG wrote:
>
> Dear all,
>
> I always use the SUPERPOSE tool in CCP4 to superpose molecules. This time I
> want to use the RMSD values of superposed C-alpha atoms to plot a RMSD graph
> (instead of using the graph automatically made by the program). However,
> there are many atoms missing in the RMSD list.
>
> In the settings I chose “Superpose specific atoms/residues”, checked “Output
> all distances to a file”, fit “C-alpha atoms”. The superposed structures
> have exactly the same sequence.
>
> My question is: is there any way to get the completed list of RMSD value for
> each C-alpha atom? Or is there any other program for this purpose?
>
> Thank you!
>
> Kind regards,
> Wenhe
>
>

Re: [ccp4bb] [Fwd: Re: [ccp4bb] FW: New ligand 3-letter code (help-7071)]

[Martyn Symmons]

Well the problem is there is a lot more to a ligand than PDB
coordinates - little things like bond orders... In addition people can
publish ligands with atoms for which they have no density - so
zero-occupancy is allowed too. So who should get priority - the group
who publishes a ligand first, or the ones who actually have density
for all the atoms?

These sorts of complications mean we all benefit from peer-review of
the structure - that is why we put things on hold. And authors should
have a chance to change their ligand definition based on reviewers'
comments - just as they are allowed to improve the PDB coordinates. So
it is a worry for them that the PDB might 'publish' the ligand aspect
of  their work before they have completed the peer-review process.

Maybe you don't believe is peer-review - in reply to which I'd
paraphrase what people say about democracy - it's pretty bad but
better than the alternatives.

But to return to the point I made: what really is the problem with
maintaining and modifying _separate_ definitions with authors'
_separate_ deposited coordinates (and bond orders) while structures
are on hold and being reviewed? Journals manage to keep all those
submitted papers separate in their databases.

cheers
 M.

On Mon, Jun 22, 2015 at 3:12 AM, Edward A. Berry ber...@upstate.edu wrote:
   I can't imagine a journal doing that can you?  When I work on my
 supplementary material in a paper I don't expect that the journal will
 take a bit out and publish it separately to support the work of my
 competitors. Not out of spite that I was beaten - but because I don't
 want to take the responsibility for checking their science for them!


 I don't see the problem here. What about the dozens of authors who
 will benefit from using your ligand in their structure _after_ your
 structure comes out? You don't take responsibility for checking their
 science. Every author gets a copy of his final structure to check
 before it is released and each is responsible for his own.
 The only difference here is whether the competitor got to use it first,
 (which might sting a bit) or only after you had already made it your
 own with the first structure.

 I guess the ligand database is the responsibility of the pdb, but
 they depend on first depositors to help set up each ligand, so
 it is not surprising if the type model has coordinates from the
 first depositor's structure (although it would be convenient if
 they were all moved to c.o.m. at 0,0,0). When another group publishes
 a structure with the ligand, they will not be publishing the first
 depositor's coordinates because the ligand will be moved to its position
 in their structure and refined against their data, probably with
 somewhat different restraints.

 If the ligand is a top secret novel drug lead that your company is
 developing I guess it would come as a shock to find someone else has
 already deposited it, and it might be good to hasten not the
 publication but protecting of the compound with a patent!

 Although Miriam says a new 3-letter code is generated when no match is
 found,
 I believe the depositor's code will be used if it is available,
 at least one of mine was last year, so there is some use for Nigel's
 utility if you want to stamp your new compound with a rememberable name.

 eab


 On 06/21/2015 06:33 PM, Martyn Symmons wrote:

 Miri raises important points about issues in the PDB Chemical
 Component Dictionary - I think part of the problem is that this is
 published completely separately from the actual PDB - so for example I
 don't think we have an archive of the CCD for comparison alongside the
 PDB snapshots? This makes it difficult to follow the convoluted track
 of particular ligands through the PDB's many,many changes to small
 molecule definitions.

 But following discussion with other contributors offline I want to
 make it clear what is my understanding of the ZA3 (2Y2I /2Y59) case:

 I am clear there was no unethical behaviour by either group in the
 course of their work on these structures and the publication of them.

 The problem I am highlighting is that the PDB don't understand
 publishing ethics - what happened in ZA3 was that they published a
 little bit of one group's work to support the work of someone who was
 scooping them!

   I can't imagine a journal doing that can you?  When I work on my
 supplementary material in a paper I don't expect that the journal will
 take a bit out and publish it separately to support the work of my
 competitors. Not out of spite that I was beaten - but because I don't
 want to take the responsibility for checking their science for them!

 All the best
Martyn

 Cambridge

 On Sun, Jun 21, 2015 at 7:01 PM, Miri Hirshberg
 02897e8e9f0f-dmarc-requ...@jiscmail.ac.uk wrote:

 Sun., June 21st 2015

 Good evening,

 adding several general points to the thread.

 (1) Fundamentally PDB unlike other chemical databases
 insists that all equal structures should have the same 3

Re: [ccp4bb] [Fwd: Re: [ccp4bb] FW: New ligand 3-letter code (help-7071)]

[Martyn Symmons]

 the same compound, 
  i.e. same 2D structure or connectivity.  Each deposition should have its 
  own 3D coordinates.  If a different publication gets your ligand 3D 
  coordinates (2Y59 actually embodies the atomic coordinates from the 
  2Y2I), that looks to me an oversight by PDB.  It is hard to believe that 
  PDB intended to use the 3D coordinates from one entry for the other, 
  ligand or not.  In fact, the restraints as described by the ligand 
  dictionary should also be kept separate as that reflects how the authors 
  refine their ligand.
 
  Yong
 
  -Original Message-
  From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of 
  Martyn Symmons
  Sent: Friday, June 19, 2015 8:39 PM
  To: CCP4BB@JISCMAIL.AC.UK
  Subject: Re: [ccp4bb] FW: New ligand 3-letter code (help-7071)
 
  By oversimplifying the situation here the PDB does not answer my related 
  point about competing crystallographers:
  My scenario:
 
  Group A deposits structure with new drug - gets their three-letter code 
  for example ZA3  they then get to check the coordinates and chemical 
  definition of this ligand.
 
  But suppose a little after that a competing group B deposits their 
  structure with the same drug which they think is novel - but no...
  they get assigned the now described ZA3 which has been checked by the 
  other group.
 
   Then it is a race to see who gets to publish and release first. And if it 
  is the second group B who wins then they are publishing the work of their 
  A competitors - who have done the depositing and checking of the ligand  
  description.
 
   Sounds unlikely? Well, it actually happened in 2011 for my exact example 
  ZA3 - present in 2Y2I and in 2Y59 from competing groups.
 
   From the dates in the mmcif it was 2Y2I depositors who set up and had a 
  chance to review the description of ZA3 ligand. Only to see it released a 
  week before their crystal structure, when their ZA3 appeared to accompany 
  competing 2Y59! It is amazing that the PDB did not spot this and arrange a 
  suitable workaround.
 
  Just to check:
  mmcif for ZA3 shows it was created for 2Y2I:
  ...
  _chem_comp.pdbx_model_coordinates_db_code2Y2I
  ...
  But it was modified for release:
  ...
  _chem_comp.pdbx_modified_date2011-07-22
  ...
  corresponding to the early 2011-07-27 release date of the competing
  structure: 2Y59 even though this PDB was  _deposited_ second.
 
  The ZA3 ligand definition released with 2Y59 actually embodies the atomic 
  coordinates from the 2Y2I structure:
 
  mmcif
  ZA3 O6   O6   O 0  1 N N N 8.279  7.165  40.963 0.311  -1.061 -0.920
  O6   ZA3 1
  ZA3 C5   C5   C 0  1 N N N 9.132  8.047  40.908 0.147  -0.205 -0.073
  C5   ZA3 2  ...
  PDB 2Y2I
  HETATM 3598  O6  ZA3 A1000   8.279   7.165  40.963  1.00 41.25 
O
  HETATM 3599  C5  ZA3 A1000   9.132   8.047  40.908  1.00 63.20
C ...
 
  Surely a better approach would be to allow both groups a chance to work 
  through and sign off on independent ligand descriptions?
 
  Then whoever releases first would release both a novel structure and the 
  ligand definition _they_ deposited and checked. Their priority can then be 
  asserted and the other group contacted to ask if they agree to accept this 
  definition. This also has the advantage of better confidentiality 
  pre-publication.
 
  Another problem from any cross-linking of definitions is that say group A 
  are motivated by reviewers' reports to change the definition of ZA3 
  pre-release. Well now the change impinges on the chemical meaning of other 
  group B's deposited structure. For example ZA3 mmcif has a statement:
 
  ZA3 Modify aromatic_flag 2011-06-04 RCSB
 
  so this change was pre-release - but we cannot be sure what motivated this 
  - whether it was signed off by the 2Y2I authors or the 2Y59 authors (or 
  both?)
 
  With the accelerating pace of drug discovery for sure this sort of 
  uncertainty is going to happen again.Unless the PDB have changed their 
  practice for ligand deposition?
 
  All the best
   Martyn
 
  Cambridge.
 
  On Fri, Jun 19, 2015 at 1:49 PM, Sheriff, Steven steven.sher...@bms.com 
  wrote:
   All:
  
  
  
   Since the original query was cross-posted on both the COOT mailing
   list and the CCP4BB Rachel Green gave me permission to forward this to
   both. She provides links about the mechanism of assignment of 3-letter
   codes. In the third link below, my original suggestion to the COOT
   mailing list that one could just use UNK is incorrect as that is 
   reserved for unknown amino acids.
   According to this document, I should have suggested UNL for an unknown
   ligand.
  
  
  
   Steven
  
  
  
   From: Rachel Kramer Green [mailto:kra...@rcsb.rutgers.edu]
   Sent: Tuesday, June 16, 2015 10:21 AM
   To: Sheriff, Steven
   Cc: info
   Subject: Re: New ligand 3-letter code (help-7071)
  
  
  
   Dear Steven,
  
   During annotation of ligands, all chemical

Re: [ccp4bb] FW: New ligand 3-letter code (help-7071)

[Martyn Symmons]

By oversimplifying the situation here the PDB does not answer my
related point about competing crystallographers:
My scenario:

Group A deposits structure with new drug - gets their three-letter
code for example ZA3
 they then get to check the coordinates and chemical definition of this ligand.

But suppose a little after that a competing group B deposits their
structure with the same drug which they think is novel - but no...
they get assigned the now described ZA3 which has been checked by the
other group.

 Then it is a race to see who gets to publish and release first. And
if it is the second group B who wins then they are publishing the work
of their A competitors - who have done the depositing and checking of
the ligand  description.

 Sounds unlikely? Well, it actually happened in 2011 for my exact
example ZA3 - present in 2Y2I and in 2Y59 from competing groups.

 From the dates in the mmcif it was 2Y2I depositors who set up and had
a chance to review the description of ZA3 ligand. Only to see it
released a week before their crystal structure, when their ZA3
appeared to accompany competing 2Y59! It is amazing that the PDB did
not spot this and arrange a suitable workaround.

Just to check:
mmcif for ZA3 shows it was created for 2Y2I:
...
_chem_comp.pdbx_model_coordinates_db_code2Y2I
...
But it was modified for release:
...
_chem_comp.pdbx_modified_date2011-07-22
...
corresponding to the early 2011-07-27 release date of the competing
structure: 2Y59 even though this PDB was  _deposited_ second.

The ZA3 ligand definition released with 2Y59 actually embodies the
atomic coordinates from the 2Y2I structure:

mmcif
ZA3 O6   O6   O 0  1 N N N 8.279  7.165  40.963 0.311  -1.061 -0.920
O6   ZA3 1
ZA3 C5   C5   C 0  1 N N N 9.132  8.047  40.908 0.147  -0.205 -0.073
C5   ZA3 2  ...
PDB 2Y2I
HETATM 3598  O6  ZA3 A1000   8.279   7.165  40.963  1.00 41.25   O
HETATM 3599  C5  ZA3 A1000   9.132   8.047  40.908  1.00 63.20
  C ...

Surely a better approach would be to allow both groups a chance to
work through and sign off on independent ligand descriptions?

Then whoever releases first would release both a novel structure and
the ligand definition _they_ deposited and checked. Their priority can
then be asserted and the other group contacted to ask if they agree to
accept this definition. This also has the advantage of better
confidentiality pre-publication.

Another problem from any cross-linking of definitions is that say
group A are motivated by reviewers' reports to change the definition
of ZA3 pre-release. Well now the change impinges on the chemical
meaning of other group B's deposited structure. For example ZA3 mmcif
has a statement:

ZA3 Modify aromatic_flag 2011-06-04 RCSB

so this change was pre-release - but we cannot be sure what motivated
this - whether it was signed off by the 2Y2I authors or the 2Y59
authors (or both?)

With the accelerating pace of drug discovery for sure this sort of
uncertainty is going to happen again.Unless the PDB have changed their
practice for ligand deposition?

All the best
 Martyn

Cambridge.

On Fri, Jun 19, 2015 at 1:49 PM, Sheriff, Steven steven.sher...@bms.com wrote:
 All:



 Since the original query was cross-posted on both the COOT mailing list and
 the CCP4BB Rachel Green gave me permission to forward this to both. She
 provides links about the mechanism of assignment of 3-letter codes. In the
 third link below, my original suggestion to the COOT mailing list that one
 could just use UNK is incorrect as that is reserved for unknown amino acids.
 According to this document, I should have suggested UNL for an unknown
 ligand.



 Steven



 From: Rachel Kramer Green [mailto:kra...@rcsb.rutgers.edu]
 Sent: Tuesday, June 16, 2015 10:21 AM
 To: Sheriff, Steven
 Cc: info
 Subject: Re: New ligand 3-letter code (help-7071)



 Dear Steven,

 During annotation of ligands, all chemical components present in the
 structure are compared against the definitions in the Chemical Component
 Dictionary (http://www.wwpdb.org/data/ccd). If the ligand is not in the
 dictionary, a three letter code is assigned. See
 http://www.wwpdb.org/documentation/policy#toc_assignment.  In the future, a
 group of three-letter codes may be set aside to be used during refinement to
 flag new ligands.

 Clarification about the ligand ids assignment and in particular the usage of
 UNX/UNL/UNK residues can be found at
 http://www.wwpdb.org/documentation/procedure#toc_2.

 Best wishes,
 Rachel



 

 Rachel Kramer Green, Ph.D.

 RCSB PDB

 kra...@rcsb.rutgers.edu



 New! Deposit X-ray data with the wwPDB at:

 http://deposit.wwpdb.org/deposition (NMR and 3DEM coming soon).

 ___

 Twitter: https://twitter.com/#!/buildmodels

 Facebook: http://www.facebook.com/RCSBPDB







 On 6/5/2015 7:50 AM, Sheriff, Steven wrote:

 All:



 Why the concern for unassigned 

Re: [ccp4bb] New ligand 3-letter code

[Martyn Symmons]

One thing to beware of is that of course un-released structures may have
the code that you choose - structures can be on hold for over a year (the
rules say a year but in practice release is often slower) and any new
ligands and codes with them. Ligand codes are assigned at deposition and
the description stored privately in a wwPDB database I believe. So your
ligand code may be already there but not visible until it is copied into
the public Chemical Component Dictionary at the time of release.

Conversely your 'new' ligand _structure_ may be in an un-released entry
deposited by some other crystallographer and already assigned a 3-letter
code. So your choice will be overwritten at deposition in any case.

This produces a problem when multiple competing groups deposit structures
containing the same novel ligand. As the database will not expect
duplicates, only the first depositor will get assigned a new 3-letter code.

However then there should be alarm bells for the second depositor if their
'novel' ligand turns out to have a pre-existing three letter code! I
suspect there must have been cases where this happened but was not spotted
by the depositors. But if it happens to you then maybe it's time to speed
up publication...

I don't know if it has happened that one crystallographer has 'hastened'
the release of the ligand description of another worker by publishing it in
complex with a solved macromolecule. Perhaps the clever people at the PDB
do a switch at the last minute?

Best wishes,
 Martyn

Martyn Symmons
Cambridge






On Mon, Jun 8, 2015 at 3:12 AM, Lau Sze Yi (SIgN) 
lau_sze...@immunol.a-star.edu.sg wrote:

 Thanks Eleanor for relaying my question from COOT and everyone that
 follows up with the thread.

 So this new ligand code search feature will be available in Phenix Gui?
 Look forward to it.

 Thanks,
 Sze Yi




 From: Nigel Moriarty nwmoria...@lbl.gov
 Reply-To: Nigel Moriarty nwmoria...@lbl.gov
 Date: Sunday, 7 June 2015 11:18 pm
 To: CCP4BB@JISCMAIL.AC.UK CCP4BB@JISCMAIL.AC.UK
 Subject: Re: [ccp4bb] New ligand 3-letter code

 Ed

 Thanks for the feature request. I will be available in the next nightly
 build.

 nigel% elbow.get_new_ligand_code A?3

 Unique ligand code : AV3

 Cheers

 Nigel

 ---
 Nigel W. Moriarty
 Building 64R0246B, Physical Biosciences Division
 Lawrence Berkeley National Laboratory
 Berkeley, CA 94720-8235
 Phone : 510-486-5709 Email : nwmoria...@lbl.gov
 Fax   : 510-486-5909   Web  : CCI.LBL.gov

 On Sat, Jun 6, 2015 at 4:20 PM, Edward A. Berry ber...@upstate.edu
 wrote:

 Neat! It's true the PDB will choose a unique code for you,
 but they will use what you supply if it is already unique,
 and it is nice to be able to choose something that can help
 you remember what it stands for.
 Gone are the days when we could choose meaningfull pdb ID's
 (like 1PRC, 2PRC, 3PRC etc are all photosynthetic reaction center),
 but there is still some freedom in choosing the ligand ID's.
 They are filling up fast, though!

 locust 123% elbow.get_new_ligand_code AC
 Unique ligand code : AC3

 locust 129% elbow.get_new_ligand_code A?3
 Unique ligand code : A?3

 eab

 On 06/06/2015 12:13 PM, Nigel Moriarty wrote:

 I wrote something a while ago the finds an unused code.

 % elbow.get_new_ligand_code


 Unique ligand code : 7V8

 % elbow.get_new_ligand_code A


 Unique ligand code : A6E


 Cheers

 Nigel

 ---
 Nigel W. Moriarty
 Building 64R0246B, Physical Biosciences Division
 Lawrence Berkeley National Laboratory
 Berkeley, CA 94720-8235
 Phone : 510-486-5709 Email : nwmoria...@lbl.gov
 Fax   : 510-486-5909   Web  : CCI.LBL.gov http://CCI.LBL.gov

 On Fri, Jun 5, 2015 at 6:59 AM, Eleanor Dodson 
 eleanor.dod...@york.ac.uk mailto:eleanor.dod...@york.ac.uk wrote:


 I use any 3 letter/number code that i want. If you read the
 corresponding cif file into coot it is used in preference to any in the
 library. The PDB deposition team will assign a code if it is a new ligand
 to the database. Could you relay this to original poster?

 Thanks

 Jim Brannigan


 On 5 June 2015 at 14:58, Eleanor Dodson eleanor.dod...@york.ac.uk
 mailto:eleanor.dod...@york.ac.uk wrote:

 OK - thank you.
 How are things?
 E


 -- Forwarded message --
 From: *Jim Brannigan* jim.branni...@york.ac.uk mailto:
 jim.branni...@york.ac.uk
 Date: 5 June 2015 at 14:39
 Subject: Re: New ligand 3-letter code
 To: Eleanor Dodson eleanor.dod...@york.ac.uk mailto:
 eleanor.dod...@york.ac.uk


 Hi Eleanor

 I use any 3 letter/number code that i want. If you read the
 corresponding cif file into coot it is used in preference to any in the
 library. The PDB deposition team will assign a code if it is a new ligand
 to the database. Could you relay this to original poster?

 Thanks

 Jim Brannigan

 On 5 June 2015 at 11:28, Eleanor Dodson 
 eleanor.dod...@york.ac.uk

Re: [ccp4bb] Antw: Re: [ccp4bb] Offtopic: Software to closely visualize interacting partnets in protein complex

[MARTYN SYMMONS]

As this is a ccp4 bb we should mention the jsPISA server which is at 
http://www.ccp4.ac.uk/pisa/
This is a very nice implementation of the PISA approach.
Another approach is to use the Chimera clash finding approach 
http://www.cgl.ucsf.edu/chimera/docs/ContributedSoftware/findclash/findclash.html
But this defaults to atom to atom contacts which means you have to prune it 
down with a script for picking residues. Also does not know about H-bond 
angles
Cheers
Martyn
Cambridge
Original message
From : bar...@fmp-berlin.de
Date : 18/03/2015 - 14:18 (GMTST)
To : CCP4BB@JISCMAIL.AC.UK
Subject : [ccp4bb] Antw: Re: [ccp4bb] Offtopic: Software to closely visualize 
interacting partnets in protein complex
Hi,
I just checked the PIC sever tim suggested. very nice indeed. If 
you only want to map different interfaces and the amino acids involved 
in, I suggest to run the pisa server, too. http://www.ebi.ac.uk/pdbe/pisa/ . I 
used it extensively to find out whether a certain set of crystal contacs leads 
to a certain crystal packing. 
best, matthias
 Tim tim.schu...@rub.de 11.03.15 18.25 Uhr 
Hi,
Molprobity is also a nice software to do this kind of analysis - if you 
use it as implemented in phenix you also get good visualization in coot.
I also asked the pymol community to create an implementation for pymol, 
but I did not follow if somebody took that up.
Also this 'protein interactions calculation' server is very neat:
http://pic.mbu.iisc.ernet.in/
/Tim
On 10/03/15 11:25, Debasish Kumar Ghosh wrote:
 Dear All,

 Apologies for this little off-topic inquiry. I want to closely visualize the 
 interacting residues in an multimeric protein complex to understand the 
 nature of interactions. Is there any good software to give this information 
 with good clarity.
 Any suggestion is highly appreciated.

 Thanks,
 Best !!!

 Debasish Kumar Ghosh

 CSIR- Junior Research Fellow (PhD Scholar)
 C/o: Dr. Akash Ranjan
 Computational and Functional Genomics Group
 Centre for DNA Fingerprinting and Diagnostics
 Hyderabad, INDIA

 Email(s): dkgh...@cdfd.org.in, dgho...@gmail.com
 Telephone: 0091-9088334375 (M), 0091-40-24749396 (Lab)
 Lab URL: 
 http://www.cdfd.org.in/labpages/computational_functional_genomics.html

[ccp4bb] [ccp4] NAGging problem?

[MARTYN SYMMONS]

For my sins I am making figures of TLR structures. These are dimeric in 
activated forms and glycosylated. So I sort of assumed that the sugars would 
inherit the chain id of the polypeptide they are attached to. But then in 2Z7X 
(TLR1/2 heterodimer) when I coloured the gycolysation to fit with the chains it 
did not look right. Looking at the LINK record again I found the following:
LINK ND2 ASN A 199 C1  NDG A 901 1555   1555  1.45  
LINK ND2 ASN A 414 C1  NAG A 911 1555   1555  1.45  
LINK ND2 ASN A 442 C1  NAG A 921 1555   1555  1.45  
LINK ND2 ASN B  51 C1  NAG A 801 1555   1555  1.46  
LINK ND2 ASN B 163 C1  NAG B 811 1555   1555  1.45  
LINK ND2 ASN B 330 C1  NAG A 821 1555   1555  1.46  
LINK ND2 ASN B 429 C1  NAG A 831 1555   1555  1.45  
So only _one_ of the NAGs bound to chain B ends up getting a B chain id! I'm 
guessing this is because in the dimers the bulk of the sugar ends up closer to 
chain A rather than the chain B which it is attached to. 
And according the the RCSB chain IDs for all bound moieties and waters are 
assigned based on their proximity (number of contacts) to the nearest 
polymer.(http://deposit.rcsb.org/depoinfo/download/dp.pdf section 4.4.1 Chain 
ID assignment ). So is this why these chains get switched? If so, I have to say 
that is crazy - surely the covalent bonds, detected when preparing the LINK 
description, should always take precedence? 
Maybe more reasonable treatment has been applied in some cases - but I would've 
hoped that there would be some consistency - and so maybe the rules need 
re-writing to remove these anomalies. 
I was only making a figure - but for sure someone will be trying to use the 
data for something more useful and quantitative. This sort of stuff will make 
their analysis difficult.
All the best
  Martyn 
Martyn Symmons
Cambridge

Re: [ccp4bb] Command line tool for RMSD calculation

[MARTYN SYMMONS]

Mentioned recently but worth mentioning again?
I have used Chimera rmsd from its own command line (find the command line under 
the 'favorites' tab) - it is scriptable.
https://www.cgl.ucsf.edu/chimera/docs/UsersGuide/midas/rmsd.html
Must have equal number of atoms matched - use selection to make that true - for 
example restricting to Calpha atoms. 
Cheers - Martyn 
Cambridge
Original message
From : c.z...@yale.edu
Date : 15/10/2014 - 21:05 (GMTDT)
To : CCP4BB@JISCMAIL.AC.UK
Subject : [ccp4bb] Command line tool for RMSD calculation
Dear all,
I am just wondering whether there is a simple command line tool for RMSD 
calculation between two aligned structures? I just cannot find a good answer 
online, and ccp4bb can always impress me.
Thank you so much in advance,
Chen

Re: [ccp4bb] Coot - way to move the center pointer to a specific x,y,z coordinate?

[MARTYN SYMMONS]

I usually add a fiducial waters to the pdb file (chain Z - remember those?) - 
or otherwise heavy atoms which will come up in a distinctive colour usually... 
Then 'go to atom' under draw menu?
Martyn
Cambridge
Original message
From : alejandro.virru...@yale.edu
Date : 09/09/2014 - 21:17 (GMTDT)
To : CCP4BB@JISCMAIL.AC.UK
Subject : [ccp4bb] Coot - way to move the center pointer to a specific x,y,z 
coordinate?
Does anyone knowhow to move the center pointer to a specific x,y,z coordinate? 
Or to place some kind of marker at a specific x,y,z coordinate?
Thanks,
Alex

Re: [ccp4bb] correlated alternate confs - validation?

[MARTYN SYMMONS]

I think there has been historically some glitch in the deposition of some 
refmac altloc waters 
So for example  4a9x from Buster is annotated correctly
REMARK   3   PROGRAM : BUSTER 2.11.2   
HETATM 9407  O   HOH A2517 -22.429   1.144 -35.218  1.00 29.73   O  
HETATM 9408  O  AHOH A2518 -18.933  -9.507 -28.753  0.50  3.00   O  
HETATM 9409  O  BHOH A2518 -20.467  -9.093 -29.900  0.50 16.03   O  
so the A and B altloc retained.
But 
for example in 4ac7 
REMARK   3   PROGRAM : REFMAC 5.6.0117 
where these two waters look like they originally were deposited as altloc waters
HETATM 6205  O   HOH A2043  -3.612  86.934 100.843  0.50 25.12   O  
ANISOU 6205  O   HOH A2043 2630   2166   4747   -153 68   1496   O  
HETATM 6206  O   HOH A2044  -3.506  85.122 100.190  0.50 22.20   O  
ANISOU 6206  O   HOH A2044 3710   2476   2248862188 16   O  
and these two
HETATM 6328  O   HOH B2073  -1.182 117.989  73.544  0.50 15.63   O  
ANISOU 6328  O   HOH B2073 2509   1684   1745408173289   O  
HETATM 6329  O   HOH B2074  -0.561 117.641  72.161  0.50 14.37   O  
ANISOU 6329  O   HOH B2074 2023   1187   2247107123   -211   O  

These waters also end up being annotated as close contacts in Remark 500 - 
which would not have happened if they had retained their altlocs.
REMARK 500 THE FOLLOWING ATOMS ARE IN CLOSE CONTACT.
...   
REMARK 500   OHOH A  2043 OHOH A  2044  1.93
REMARK 500   OHOH B  2073 OHOH B  2074  1.56

Probably other examples can be retrieved but this latter file dates from my 
time at the PDBe and so may have been mangled at my hands.

In my defence I think it is owing to the annotation step when the waters are 
renumbered to fit with the associated macromolecule N to C (for proteins).  I 
think the script that was originally developed for this was not able to deal 
with waters with the same number but differing altlocs. Also alternate 
positions in this step can be associated with different residues by reason of 
being closer to one or other bit of chain. In this case they can have very 
different numbers - or chains.

I think this can be handled better in the new mmCif-based annotation system 
with the cooperation of refinement program authors. 
all the best
Martyn 

Martyn Symmons
Cambridge


Original message
From : f...@bernstein-plus-sons.com
Date : 23/07/2014 - 16:20 (GMTDT)
To : CCP4BB@JISCMAIL.AC.UK
Subject : Re: [ccp4bb] correlated alternate confs - validation?

I agree that it would be excellent to be able to associate
alternate conformations (beyond the individual residue) but
when we defined the PDB format we had an 80-column limitation
per atom and so only one column was allowed for alternate
conformations.  In an ASCII world only 36 characters were available
to define alternate conformations.  This is inadequate to allow
for many residues with independent alternate conformations -
one residue with three conformations would use up 3 of the 36
characters.  Thus there was no way to say that alternate
conformation A in one residue is or is not associated with
alternate conformation A in another residue.

  Frances

=
Bernstein + Sons
*   *   Information Systems Consultants
5 Brewster Lane, Bellport, NY 11713-2803
*   * ***
 *Frances C. Bernstein
   *   ***  f...@bernstein-plus-sons.com
  *** *
   *   *** 1-631-286-1339FAX: 1-631-286-1999
=

On Wed, 23 Jul 2014, Tim Gruene wrote:

 -BEGIN PGP SIGNED MESSAGE-
 Hash: SHA1

 Hi Bernhard,

 That's right, the definition is not in the PDB, but REFMAC sensibly
 handles it this way. It is certainly a short-coming if not bug in the
 PDB (definition).

 Best,
 Tim

 On 07/23/2014 04:49 PM, Bernhard Rupp wrote:
 ?? Refmac knows because of the group definition, otherwise I cannot
 force grouped occupancy refinement. There is no definition in PDB
 that eg ALTLOC A (of whatever residue) belongs to ALTLOC A of
 (whatever) residue/water.

 BR

 -Original Message- From: Tim Gruene
 [mailto:t...@shelx.uni-ac.gwdg.de] Sent: Mittwoch, 23. Juli 2014
 15:38 To: b...@hofkristallamt.org; CCP4BB@JISCMAIL.AC.UK Subject: Re:
 [ccp4bb] correlated alternate confs - validation?

 Hi Bernhard,

 do you refer to the PDB who, according to Martyn, remove the altloc
 indicator? That's certainly a serious bug that should be fixed as
 quickly as possible.

 Within refmac the preservation is fine and as you would expect it
 to be: altA only sees atoms in altA and those witout altLoc, etc,
 which makes sure you PDB file is interpreted correctly by refmac5.
 That's how I refmac works.

 Best, Tim

 On 07/23/2014 03:26 PM, Bernhard Rupp wrote:
 I would

Re: [ccp4bb] EDSTATS for an extracted fragment

[MARTYN SYMMONS]

Is n't the danger here that you are just fitting to noise generated by the rest 
of the Unit Cell that you are ignoring. So it is easy to do a 'good bit' of 
modelling in a specific region. Just like some tennis players have very good 
serves but do not win matches. 



 From: George Devaniranjan devaniran...@gmail.com
To: CCP4BB@JISCMAIL.AC.UK 
Sent: Wednesday, 11 June 2014, 19:22
Subject: Re: [ccp4bb] EDSTATS for an extracted fragment
 


Thank you Gerard for your suggestion.

I will look into the paper you suggested and MAPMAN too.

I am idealizing fragments of PDB and while this would not agree when compared 
with the  well  refined structure, my goal really is seeing if these ideal 
fragments can still be identifiable when I look at the electron density and how 
to quantify that.
(Ideal here refers to a small number of Phi and Psi values I picked as 
representative of different regions in the Ramachandran map).


Isn't the danger here that you are can't tell how much improvement is just 
fitting to noise from improvements that should be made in the rest of the 
model? 

Unless you go on and do a final full refinement you can't know if your 
modelling is warranted by a global improvement in fit.

It could be the case that you are improving globally but I don't know how you 
can tell that by fitting to just a bit of the map. 

We have all had that experience of rebuilding a residue so it looks 'right' to 
us locally but then the refinement program knows better after making a global 
assessment and puts it back. 

If some way to give local density 'error bars' that might be helpful so that a 
maximum likelihood approach could be used during real space fitting. Is that 
similar to what those useful LLG maps give in heavy atom model refinement?


regards
Martyn

Cambridge 

 



   

I picked on EDSTATS to explore since it gave individual residue information but 
realize now that since I am dealing with a small sub-set of residues an 
alternative is required.






On Wed, Jun 11, 2014 at 2:01 PM, Gerard DVD Kleywegt ger...@xray.bmc.uu.se 
wrote:

What you want is a test for how well each model agrees with its own map. It is 
fair to argue that the model that is more self-consistent (agrees better with 
its own map) is the better model.  But you won't learn that by comparing model 
A to map B.


However, conversely, if your modified model fits the original map better than 
the model that was used to calculate the map itself, you've done a good bit of 
model building. If you want to do this calculation (with all the warnings and 
caveats), you can also use MAPMAN - 
http://xray.bmc.uu.se/usf/mapman_man.html#S41 . The method you propose is 
essentially the same as this one: http://www.ncbi.nlm.nih.gov/pubmed/18598022 
but for a fragment of your macromolecule instead of for a ligand (if you don't 
have access to the journal, you can request a reprint here: 
http://xray.bmc.uu.se/cgi-bin/gerard/reprint_mailer.pl?pref=87 )

--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 !
**

Re: [ccp4bb] experimental phasing at low resolution

[MARTYN SYMMONS]

Don't forget that your SeMet has real differences from your native - the S to 
Se brings in 18 extra electrons (less obviously if there is lower 
incorporation). But that mean if your native is isomorphous enough (you didn't 
say) then you possibly have 'ordinary' isomorphous differences to work with. 
You could then work in the anomalous as a sort of SIRAS solution.  
SeMet as isomorphous has been used previously in many structure solutions.
All the best
  Martyn

Martyn Symmons
Cambridge 



 From: vijay srivastava vijaytec...@yahoo.co.in
To: CCP4BB@JISCMAIL.AC.UK 
Sent: Tuesday, 3 June 2014, 9:37
Subject: [ccp4bb] experimental phasing at low resolution
 


Dear All,
Does anyone know of a facility
that provides amino acid analysis for determination of SeMet
incorporation in a recombinant protein?
We're looking for some advice
about how to proceed with a structure we're working on.  Our
protein is 350 amino acids (10 methionine are present) and naturally binds 
magnesium.  We
have a SeMet data set at peak that goes down to  5.4 angstroms. 
We tried to find the heavy atom position with Shelxcde program. The
log file is given as below.

Resl.   Inf - 8.0 - 6.0 - 5.6 - 5.4 -
5.2 - 5.0 - 4.8 - 4.6 - 4.4 - 4.2 - 3.98 
N(data) 529   643   24129
0 0 0 0 0 0 0 
I/sig63.5  19.5  10.6 
10.0   0.0   0.0   0.0   0.0   0.0   0.0   0.0 
%Complete  94.1  98.9  99.2  18.0  
0.0   0.0   0.0   0.0   0.0   0.0   0.0 
d/sig   3.06  1.24  1.07 1.21 

I am also attaching the .lst and .res file and
from that we were considering that only one selenomethionine
position.We have one monomer per AU
(-unfortunately, MR is not working for this project). The space group
is P3 .  We also have a native set down to 3.4 angstroms. 
While we understand that we may need more phasing information  we're
wondering if anyone might have some other suggestions or insights
about how we can move forward given the data that we currently have. 
Can we extend the phases with our native data set. Thanks in advance
for any advice.

regards

 
Vijay

Re: [ccp4bb] Sea Urchin like crystals

[MARTYN SYMMONS]

Dear V.
  are you sure these are crystalline? - I have had similar growth from 
'spherulites' that I thought was promising. But then when I went in with a 
needle to break them for seeding, it turned out they were fibrous not 
crystalline. This made me think they were a product of denatured protein. So I 
would say poke them first to check they snap like a crystal would. Actually 
mine were birefringent in four big sectors similar to the domains in the 
underlying spherulites (which had a four sectors with opposite quarters the 
same colour - like the Burundi flag (had to look that up;) ) .  My feeling is 
fibrous protein is not a good crystallization lead.
  But maybe you have had better luck 
  Martyn 
  



 From: venkatareddy dadireddy venkatda...@gmail.com
To: CCP4BB@JISCMAIL.AC.UK 
Sent: Wednesday, 21 May 2014, 10:58
Subject: [ccp4bb] Sea Urchin like crystals
 


Dear All,

I'm working on 24.3 KDa protein (pI 4.84) with (Asp + Glu): 36 and (Arg + Lys): 
16. Protein is in buffer Tris (10mM), pH 7.5. I got sea urchin like crystal 
clusters @ protein conc 38.5 mg/ml in sitting drop method; condition 0.2M NH4I, 
20% PEG3350  pH 6.8 (PEG/ION A12). These crystals are very thin. I tried 
different pH values ranging from  6.0 -9.0 with 0.5 units increment, PEG conc. 
from 10 -30% and NH4I conc. from 0.1 -0.5 M independently in hanging drop 
method and ended up with NO Crystals. Can somebody suggest strategy to improve 
crystals. Please find the crystal pic with attachment.

Regards,
Venkat.

Re: [ccp4bb] PDB passes 100,000 structure milestone

[MARTYN SYMMONS]

I agree some forum for community annotation and commenting would be a good 
thing for users of structural data. 
There was an attempt to do that with the pdbwiki project which was a community 
resource for the bioinformatics community. Unfortunately pdbwiki has now folded 
(see http://pdbwiki.org/) They are now directing people to Proteopedia. However 
Proteopedia has a more educative focus I think - rather than capturing 
technical questions and input.

Pubmed commons (http://www.ncbi.nlm.nih.gov/pubmedcommons/), which is a forum 
for discussing the literature, is currently under testing. Perhaps this is the 
sort of thing that could work for structural data?
 
cheers
 Martyn 



 From: Ethan A Merritt merr...@u.washington.edu
To: CCP4BB@JISCMAIL.AC.UK 
Sent: Wednesday, 14 May 2014, 19:22
Subject: Re: [ccp4bb] PDB passes 100,000 structure milestone
 

On Wednesday, 14 May, 2014 13:52:02 Phil Jeffrey wrote:
 As long as it's just a Technical Comments section - an obvious concern 
 would be the signal/noise in the comments themselves.  I'm sure PDB 
 would not relish having to moderate that lot.
 
 Alternatively PDB can overtly link to papers that discuss technical 
 issues that reference the particular structure - wrong or fraudulent 
 structures are often associated with refereed publications that point 
 that out, and structures with significant errors often show up in that 
 way too.  I once did a journal club on Muller (2013) Acta Cryst 
 F69:1071-1076 and wish that could be associated with the relevant PDB 
 file(s).

Perhaps some combination of those two ideas?

The PDB could associate with each deposited structure  a crowd-sourced
list of published articles citing it.     They already make an effort to
attach the primary citation, but so far as I know there is currently
no effort to track subsequent citations.  

While spam comments in a free-format forum are probably inevitable,
spam submission of citing papers seems less likely to be a problem.

    - Ethan

  On Wed, May 14, 2014 at 12:32 PM, Zachary Wood z...@bmb.uga.edu
  mailto:z...@bmb.uga.edu wrote:
 
      Hello All,
 
      Instead of placing the additional burden of policing on the good
      people at the PDB, perhaps the entry page for each structure could
      contain a comments section. Then the community could point out
      serious concerns for the less informed users. At least that will
      give users some warning in the case of particularly worrisome
      structures. The authors of course could still reply to defend their
      structure, and it may encourage some people to even correct their
      errors.
 
-- 
Ethan A Merritt
Biomolecular Structure Center,  K-428 Health Sciences Bldg
MS 357742,   University of Washington, Seattle 98195-7742

Re: [ccp4bb] PDB passes 100,000 structure milestone

[MARTYN SYMMONS]

Dear Zachary
I once suggested this sort of discussion forum to a PDB PI as a possible 
service that could be a ' validation fight-club '  - which suggestion was not 
well received ;) But as you say it is down to setting the correct professional 
tone. 

One thing that would allow a visual discussion would be the possibility to 
share molecular representations - similar to the way Proteopedia uses Jmol 
scripting to drive the graphics from the webpages. In this way commentators 
could share viewpoint and representations to make their points. This would make 
it most useful to the wider biological community. 

all the best
 Martyn 

  



 From: Zachary Wood z...@bmb.uga.edu
To: CCP4BB@JISCMAIL.AC.UK 
Sent: Thursday, 15 May 2014, 14:47
Subject: Re: [ccp4bb] PDB passes 100,000 structure milestone
 



Adding to Tim’s comment, I would not expect a tremendous amount of spurious 
comments about a single PDB out of 100,000 unless there was a problem. 
Especially if the Pubmed Commons model was applied, and only depositors could 
comment. I would assume this would be very beneficial, given that we are 
conscientious professionals.  Could actually be a great forum for authors to go 
a little deeper into specific approaches or problems that they had with a 
structure. Not all interesting details make it to the pub.  



Best regards,

Z


***
Zachary A. Wood, Ph.D.
Associate Professor                      
Department of Biochemistry  Molecular Biology
University of Georgia
Life Sciences Building, Rm A426B
120 Green Street
Athens, GA  30602-7229
Office: 706-583-0304
Lab:    706-583-0303
FAX: 706-542-1738
***







On May 15, 2014, at 9:39 AM, Tim Gruene t...@shelx.uni-ac.gwdg.de wrote:

-BEGIN PGP SIGNED MESSAGE-
Hash: SHA1

Dear all,

isn't the ccp4bb a very good example that spam may not be such an
issue for a discussion platform on structures in the PDB? There is a
great variety of opinions, some to agree with, some to disagree, but
all of them interesting and contributing, and I hardly remember a
message I would classify as spam. And it all works without restraints.

Best,
Tim


On 05/15/2014 03:21 PM, Zachary Wood wrote:

I agree with Martyn,

Pubmed Commons could be a great model. I believe you have to be a
published author to obtain an account. It might cut down on some of
the spam/noise if the PDB adopted such a model for depositors.

Best regards,

Z


*** Zachary A. Wood,
Ph.D. Associate Professor Department of Biochemistry  Molecular
Biology University of Georgia Life Sciences Building, Rm A426B 120
Green Street Athens, GA  30602-7229 Office: 706-583-0304 Lab:
706-583-0303 FAX: 706-542-1738 
***







On May 15, 2014, at 7:29 AM, MARTYN SYMMONS
martainn_oshioma...@btinternet.com wrote:


I agree some forum for community annotation and commenting would
be a good thing for users of structural data. There was an
attempt to do that with the pdbwiki project which was a community
resource for the bioinformatics community. Unfortunately pdbwiki
has now folded (see http://pdbwiki.org/) They are now directing
people to Proteopedia. However Proteopedia has a more educative
focus I think - rather than capturing technical questions and
input.

Pubmed commons (http://www.ncbi.nlm.nih.gov/pubmedcommons/),
which is a forum for discussing the literature, is currently
under testing. Perhaps this is the sort of thing that could work
for structural data?

cheers Martyn

From: Ethan A Merritt merr...@u.washington.edu To:
CCP4BB@JISCMAIL.AC.UK Sent: Wednesday, 14 May 2014, 19:22 
Subject: Re: [ccp4bb] PDB passes 100,000 structure milestone

On Wednesday, 14 May, 2014 13:52:02 Phil Jeffrey wrote:

As long as it's just a Technical Comments section - an obvious
concern would be the signal/noise in the comments themselves.
I'm sure PDB would not relish having to moderate that lot.

Alternatively PDB can overtly link to papers that discuss
technical issues that reference the particular structure -
wrong or fraudulent structures are often associated with
refereed publications that point that out, and structures with
significant errors often show up in that way too.  I once did a
journal club on Muller (2013) Acta Cryst F69:1071-1076 and wish
that could be associated with the relevant PDB file(s).

Perhaps some combination of those two ideas?

The PDB could associate with each deposited structure  a
crowd-sourced list of published articles citing it.    They
already make an effort to attach the primary citation, but so far
as I know there is currently no effort to track subsequent
citations.

While spam comments in a free-format forum are probably
inevitable, spam submission of citing papers seems less likely to
be a problem.

- Ethan


On Wed, May 14, 2014 at 12:32 PM, Zachary Wood
z...@bmb.uga.edu mailto:z

Re: [ccp4bb] Renumbering of water molecules during deposition

[MARTYN SYMMONS]

I had thought that after discussion on this bulletin board we could find no 
rational scientific reason why the PDB renumbers waters from the N to the C 
term of the polypeptides (and 5' to 3' of nucleic chains?). 
https://www.jiscmail.ac.uk/cgi-bin/webadmin?A2=ind1311L=CCP4BBD=0P=12314

I think that provided you follow the agreed rules on chain ids (see below) then 
you have a 'right of veto' on the water renumbering. You also have a 'right of 
veto' for the water movement to symmetry-related positions - which would 
involve chain id change. 

So probably the best way is to assign the chain ids yourself and just say you 
don't want any of your waters renumbered. Then keep track of them yourself as 
Ed suggested using documented community tools.

One related question is how waters with alternate positions are dealt with. I 
cannot find clear documentation on this. Often sidechains with alternate 
positions motivate alternate waters during refinement. But I believe these are 
removed during the deposition water renumbering step. So if you have particular 
waters in alternative positions - say owing to a Michaelis complex at the 
active site - then the renumbering will likely split your water in alternate 
positions to produce two apparently unrelated, differently-numbered, solvent 
atoms. It could be argued how can you know these are alt pos of the same water? 
But if you have refined them like that then it is good for users of your 
coordinates to see that documented.  

It may be that the new PDB Deposition System is more flexible in this regard - 
I have not used it yet.

Below is the current agreed procedure from the wwPDB website.

Hope that helps. 
 regards
  Martyn 
How are chain IDs assigned to chemical groups and waters?
All chemical groups and waters within 5 Ångstroms of any atom of a polymeric 
chain will be considered to be associated with that chain and will be assigned 
the same chain ID as that polymer. If a particular chemical group/water is 
equidistant from more than one chain, then the chain ID is randomly assigned to 
be that of any one of these polymers.
Waters further than 5 Ångstroms away from the polymer that can be moved by 
symmetry to within 5 Ångstroms, will then be automatically moved and the author 
will be notified. If there are objections, the waters will be moved back to 
their original positions.
Waters further than 5 Ångstroms away from any polymer, which cannot be brought 
closer to a polymer chain by application of symmetry, will be brought to the 
attention of the depositor. The waters will be listed in REMARK 525 with the 
distance to the nearest macromolecule listed. Authors will be given the 
opportunity to remove the waters or update the coordinates. Authors may choose 
to retain the distant waters in the entry.



 From: Edward A. Berry ber...@upstate.edu
To: CCP4BB@JISCMAIL.AC.UK 
Sent: Monday, 7 April 2014, 15:25
Subject: Re: [ccp4bb] Renumbering of water molecules during deposition
 

After deposition (my experience is with RCSB) you will get back a processed
file to verify.  See how the waters have been renumbered, and renumber
in such a a way that your original scheme is easily identifiable but in
keeping with whatever the renumbering was trying to achieve- e.g. your
waters X1,X2,X3 could become A1001, A1002, A1003 and the symmetry related
waters B1001, B1002, B1003. Discuss with the annotator to see how your
waters can be renumbered in a recognizable way in keeping with requirements
of the PDB conventions. Then in future depositions keep this same scheme
(you may still need to renumber the original processed file each time).
eab

Eckhard Hofmann wrote:
 Hi all,
 during deposition of structures the water numbering is frequently changed.
 I would love to keep specific numbers for critical water molecules e.g.
 in an active site in related proteins (mutants, ligand soaks), to
 facilitate easier discussion in a manuscript.

 Is there a general agreement how to treat these?

 One way to deal with this is would be to use an arbitrary offset:
 A1-A233 protein
 A301 ligand
 A302 another ligand
 A401-404 4 special waters
 A500- general solvent

 Any input welcome ;-)
 All the best,
 Eckhard
 --
 Prof. Dr. Eckhard Hofmann eckhard.hofm...@bph.ruhr-uni-bochum.de
 Ruhr-Uni Bochum
 AG Proteinkristallographie, LS Biophysik, ND04/318
 44780 Bochum
 Tel: +49-(0)234/32-24463, Sekr. -24461, FAX: -14762

Re: [ccp4bb] Convert DSSP to PDB format

[MARTYN SYMMONS]

The PDB don't seem to use DSSP for the PDB format coordinate files - but 
instead some sort of in-house algorithm that gives slightly different answers 
at times. 

This can trip you up - I recently loaded 3mop into Pymol and then deleted the 
header including the PDB secondary structure and loaded it again. Surprisingly 
deleting the PDB header gave  significant differences in the death domain 
helices  - most likely because Pymol reads the PDB secondary structure if 
present; or otherwise follows its own version of DSSP rules.   

To add to confusion the PDB actually shows the DSSP-based secondary structure 
for the chains in 3mop 
(http://www.rcsb.org/pdb/explore/remediatedSequence.do?structureId=3MOPbionumber=1)
 and apparently other structures. 


You might expect that the secondary structure in the files downloaded from the 
PDB would correspond to that displayed on their own webpages - but not so 

I don't know of a publication describing the secondary structure dictionary 
used by the PDB deposition software. But if you want to be sure of what appears 
in the header of your deposited file then best thing is to use dssp2pdb as 
suggested.  Then supply the output to the PDB. They will replace their version 
with the ones you provide. But will include remarks in the PDB files to 
indicate that you have not followed their standard procedure (for example: 
REMARK 650 DETERMINATION METHOD: AUTHOR PROVIDED.).

I don't know if the dictionary used by the new mmCif-based Deposition and 
Annotation system will produce definitions equivalent to those used previously 
to produce the PDB file versions. Or if the new DA now produces definitions 
that are equivalent to DSSP. Or if it produces some third type of definition... 

Perhaps someone at the PDB will clarify - when I ask them specific questions as 
an individual then they don't bother replying - but if the bulletin board shows 
sufficient interest then we usually get at 'Public Service Announcement' of 
some kind ;) 

All the best
  Martyn 

Martyn Symmons
Cambridge



 From: Mark Brooks mark.x.bro...@gmail.com
To: CCP4BB@JISCMAIL.AC.UK 
Sent: Saturday, 1 March 2014, 16:19
Subject: Re: [ccp4bb] Convert DSSP to PDB format
 

Hi Wenhe,
       One answer to Q#2 is James Stroud's dssp2pdb to take the
output of DSSP  write HELIX and SHEET lines etc.
http://www.jamesstroud.com/software/dssp2pdb/

Mark
P.S. I believe the answer to question #1 is DSSP but I suspect the
answer is more complex.

On 1 March 2014 13:50, Wenhe wenhezhong.xmu@gmail.com wrote:
 Dear CCP4 friends,

 I have two questions after I solving one structure:

 1. What tool does PDB databank use to generate secondary structure 
 information which also shows in the pdb. file?
 2. I had a secondary structure file which generated by DSSP, how I can 
 convert it to pdb format?

 Thank you.

 Kind regards,
 Wenhe

Re: [ccp4bb] resubmission of pdb

[MARTYN SYMMONS]

Yes, thanks Robbie
That is just my point - a structure submitted and then validated for paper 
reviewers by the PDB can be changed almost completely after the paper is 
accepted.
The data can be changed too and only stipulation seems to be that it cannot be 
new data i.e. it has to have a date of collection _before_ submission. 

Changes are of course not bad in principle - they may be motivated by peer 
review.

But they need to be tracked. ln a way that is understandable for users of the 
data. Perhaps they are available in the new mmCif-based deposition and 
annotation system. I have not used it yet. 

The PDB provides a comparison between obsoleted and superseding entries 
(coordinates at least). So a similar approach could be used.

all the best
Martyn

 


 From: Robbie Joosten robbie_joos...@hotmail.com
To: CCP4BB@JISCMAIL.AC.UK 
Sent: Sunday, 2 February 2014, 20:13
Subject: Re: [ccp4bb] resubmission of pdb
  

Hi Martyn,

 I have recently had the same problem. But generally, the PDB will usually
 allow a further 6 months hold for review or modifications to an already
 submitted paper.
That is good to hear. I guess the 1 year limit is mostly to avoid structures
to stay in limbo too long.

 But what I wanted to say was that the correct term is 'withdrawal' if the
entry
 is removed pre-release - 'retraction' carries a pejorative connotation.
Even
 after release, pulling an entry would be called obsoleting (status OBS)
 without superseding. So some structures have been 'obsoleted' owing to
 retraction of a published paper. (Superseding is when a better structure
 replaces the original - this process is tracked by the PDB.)
Indeed, bad choice of words on my side. Just to complete the list, the
possible statuses are here: http://www.ebi.ac.uk/pdbe-srv/status/search/doc

 Most pre-release 'withdrawn' entries are of course subsequently released
 after re-submission. But the PDB does not seem to track these connections
-
 although they maintain a list of withdrawn entries - which means ids
cannot
 really be recycled.
That's too bad, there are not that many possible PDBids.

 Interestingly, before release entries can be 'replaced' which means a new
 structure can take the place (and 4 letter code) of the old one - this
would
 have to have the same meta-data - so source and expression - but could
 have different resolution, space group, coordinates, and small molecules.
 Changes in these could for example be motivated by referees' comments on
 the submitted paper or maybe the authors got lucky with a better crystal.
But
 this pre-release replacement could also be potentially used to 'sex up' a
 structure - for example by adding a 'novel' small molecule 'overlooked' in
the
 original deposition. Such changes are tracked privately by the PDB but are
not
 publically available... even after release.
I didn't know this was an option. It seems sensible for peer review, but
does present a potential loop-hole. I saw a fairly recent PDB entry that was
deposited as a C-alpha trace (in 2013), but presented as a full model in
Table 2 of the linked publication. The model was deposited a month before
the paper was accepted, so referees could have noticed this (in theory). But
now I wonder I the model was not 'downgraded' before the release. Perhaps
I'm just paranoid.

Cheers,
Robbie

 Even more interestingly, the ligand definitions such as bond orders can be
 modified _after_ release (as in the recent R12 case I noticed*)... I think
this is
 owing to the lack of clear rules on small molecule changes - which means
the
 PDB should be considered of limited value as a definitive record of small
 molecule chemistry.
 
 Cheers - M
 *https://www.mail-archive.com/ccp4bb@jiscmail.ac.uk/msg33403.html
 
 
 
 
 
 
 
 
 From: Robbie Joosten robbie_joos...@hotmail.com
 To: CCP4BB@JISCMAIL.AC.UK
 Sent: Saturday, 1 February 2014, 12:48
 Subject: Re: [ccp4bb] resubmission of pdb
 
 
 Hi Folmer,
 
 Perhaps because of the one year limit of keeping PDB entries in the 'HPUB'
 status.
 
 So when a PDB entry is retracted before release, is the PDBid recycled
after
 a while?
 
 Cheers,
 Robbie
 
  -Original Message-
  From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of
  Folmer Fredslund
  Sent: Saturday, February 01, 2014 10:33
  To: CCP4BB@JISCMAIL.AC.UK
  Subject: Re: [ccp4bb] resubmission of pdb
 
  Hi Faisal,
 
  There is one thing I don't understand:
 
  Some time back i had submitted a coordinate in PDB but because of
  unacceptance of the manuscript we had to retract the submission
 
  Why would you need to retract your deposited structure just because the
  paper describing the structure didn't get accepted?
 
 
  Venlig hilsen
  Folmer Fredslund
 
  On Jan 31, 2014 10:04 PM, Faisal Tarique faisaltari...@gmail.com
 wrote:
 
 
      Dear all
 
      Dear Dr. PDB,
 
      Some time back i had submitted a coordinate in PDB but because of
  unacceptance of the manuscript we had to 

Re: [ccp4bb] resubmission of pdb

[MARTYN SYMMONS]

I have recently had the same problem. But generally, the PDB will usually allow 
a further 6 months hold for review or modifications to an already submitted 
paper. 

But what I wanted to say was that the correct term is 'withdrawal' if the entry 
is removed pre-release - 'retraction' carries a pejorative connotation. Even 
after release, pulling an entry would be called obsoleting (status OBS) without 
superseding. So some structures have been 'obsoleted' owing to retraction of a 
published paper. (Superseding is when a better structure replaces the original 
- this process is tracked by the PDB.)

Most pre-release 'withdrawn' entries are of course subsequently released after 
re-submission. But the PDB does not seem to track these connections - although 
they maintain a list of withdrawn entries - which means ids cannot really be 
recycled. 

Interestingly, before release entries can be 'replaced' which means a new 
structure can take the place (and 4 letter code) of the old one - this would 
have to have the same meta-data - so source and expression - but could have 
different resolution, space group, coordinates, and small molecules. Changes in 
these could for example be motivated by referees' comments on the submitted 
paper or maybe the authors got lucky with a better crystal. But this 
pre-release replacement could also be potentially used to 'sex up' a structure 
- for example by adding a 'novel' small molecule 'overlooked' in the original 
deposition. Such changes are tracked privately by the PDB but are not 
publically available... even after release.

Even more interestingly, the ligand definitions such as bond orders can be 
modified _after_ release (as in the recent R12 case I noticed*)... I think this 
is owing to the lack of clear rules on small molecule changes - which means the 
PDB should be considered of limited value as a definitive record of small 
molecule chemistry.

Cheers - M
*https://www.mail-archive.com/ccp4bb@jiscmail.ac.uk/msg33403.html
 






 


 From: Robbie Joosten robbie_joos...@hotmail.com
To: CCP4BB@JISCMAIL.AC.UK 
Sent: Saturday, 1 February 2014, 12:48
Subject: Re: [ccp4bb] resubmission of pdb
  

Hi Folmer,

Perhaps because of the one year limit of keeping PDB entries in the 'HPUB'
status. 

So when a PDB entry is retracted before release, is the PDBid recycled after
a while? 

Cheers,
Robbie

 -Original Message-
 From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of
 Folmer Fredslund
 Sent: Saturday, February 01, 2014 10:33
 To: CCP4BB@JISCMAIL.AC.UK
 Subject: Re: [ccp4bb] resubmission of pdb
 
 Hi Faisal,
 
 There is one thing I don't understand:
 
 Some time back i had submitted a coordinate in PDB but because of
 unacceptance of the manuscript we had to retract the submission
 
 Why would you need to retract your deposited structure just because the
 paper describing the structure didn't get accepted?
 
 
 Venlig hilsen
 Folmer Fredslund
 
 On Jan 31, 2014 10:04 PM, Faisal Tarique faisaltari...@gmail.com
wrote:
 
 
     Dear all
 
     Dear Dr. PDB,
 
     Some time back i had submitted a coordinate in PDB but because of
 unacceptance of the manuscript we had to retract the submission. During
 this procedure i got few zipped file from the annotator such as 1.
 rcsb0.cif-public.gz,  2. rcsb0.pdb.gz and  3. rcsb0-
 sf.cif.gz..Now i want to submit the same ..My question is what is the best
 way to do it again..??
     Should we start  from the beginning through ADIT Deposition tool
 and resubmit it with a new PDB id or there is some way to submit again
those
 zip files which the annotator sent us after retraction..May you please
suggest
 what could be the easiest way to submit our structure to PDB without much
 efforts.
 
 
     --
     Regards
 
     Faisal
     School of Life Sciences
     JNU
 
 

Re: [ccp4bb] How to show the ligands (both as sticks and sphere) as shown here in Pymol

[MARTYN SYMMONS]

Not a pymol approach but I've noticed this dual mode display with transparency 
is the default in Molsoft ICM browser if you toggle on  ball and sticks 
together with spacefill for the same atoms.
Molsoft ball-and-stick also shows the bond order or aromaticity of ligand atoms 
- which I guess is a motivation for combining sticks with spacefill. Can export 
higher res png of the graphics too. 
  all the best
   Martyn   



 From: Wei Shi wei.shi...@gmail.com
To: CCP4BB@JISCMAIL.AC.UK 
Sent: Monday, 20 January 2014, 4:40
Subject: [ccp4bb] How to show the ligands (both as sticks and sphere) as shown 
here in Pymol
 


Hi all,

Please see attached Fig where they show the ligand both as sticks and spheres 
at 
the same time. Does anyone happen to know how to display the ligand like this? 
Thank you so much! 


Best,
Wei 

Re: [ccp4bb] Problematic PDBs

[MARTYN SYMMONS]

Hmm
So the backstory for the problematic ligand R12 in this thread is that a 
sharp-eyed worker at the wwPDB recently spotted that there was an error 
compared with the original 1999 paper. Correcting the R12 ligand is a friendly 
gesture from the PDB as it appears that the error must have been the authors' - 
the atom correcting the R12 ligand has been inserted by the PDB staff rather 
than retreived from a deposited structure.
 
It is a shame the same helpful approach is not always applied. One current 
example I spotted is a separate 'problematic ligand' 5AX which has been added 
by the wwPDB to at least four other authors' entries, starting in 2006 with the 
latest in 2009. 
 
5AX is basically a fragment ligand which the PDB software produces if a NAG has 
wandered too far from its Asn sidechain during refinement. If 5AX is generated 
during the PDB processing of a deposition, then it really should be highlighted 
for the authors as a geometric issue - rather than, as in these cases, being 
simply added to the coordinates. 
 
Reading the authors' papers for the 5AX-containing entries makes it clear that 
they never expected anything other than NAG to appear in their deposited 
coordinates.
 
And given its artifactual production during deposition, 5AX should never have 
'escaped into the wild'. 
 
So if a retrospective fix can be applied to R12 (which similar in lacking an 
atom) then it seems to me that, in fairness, a clean up of the 5AX entries 
should be arranged. 
 
Yours (not holding his breath),
Martyn
 
 

  


 From: Rachel Kramer Green kra...@rcsb.rutgers.edu
To: CCP4BB@JISCMAIL.AC.UK 
Sent: Wednesday, 6 November 2013, 16:49
Subject: Re: [ccp4bb] Problematic PDBs
  


Dear Martyn,

wwPDB staff regularly reviews and remediates PDB data and related
  dictionaries such as the Chemical Component Dictionary (CCD).

As part of our on-going remediation efforts, the chemical
  components in the archive are regularly reviewed to ensure the
  correctness and the completeness of the chemical representation.
  Such reviews show that in some cases, the author has failed to
  provide a complete description of the chemistry. To address any
  such errors, the definitions are corrected. The chemical name and
  formula are changed in the PDB file, but the coordinates are not
  changed.

In the case of entry 3CBS, issues were found with the chemical
  component definition for its ligand R12. The methyl group was not
  in the deposited coordinates and it was missing from the original
  definition. In addition, the bond order in one of the
  carbon-carbon bonds was incorrectly defined. The CCD definition
  for R12 was updated in 2011 to add the methyl group and to correct
  the bond order based on information in the primary citation. The
  coordinates for this PDB entry were not changed. Therefore, in
  accordance with wwPDB policy, the file was not obsoleted.

Sincerely,
Rachel Green


 


  
Rachel Kramer Green, Ph.D. 
RCSB PDB 
kra...@rcsb.rutgers.edu 
  
  
Twitter: https://twitter.com/#!/buildmodels 
Facebook: http://www.facebook.com/RCSBPDB 
   
On 10/21/2013 6:28 AM, MARTYN SYMMONS wrote:
 
As a postscript it might be worth mentioning one problematic ligand that 
suggested to me a way to correct some of the errors mentioned in this thread 

R12 is indicated as 9-(4-HYDROXY-2,6-DIMETHYL-PHENYL)-3 in the  most 
recent Coot monomer library. But in the PDB ligand description it is 
9-(4-hydroxy-2,3,6-trimethylphenyl)-3,7-dimethylnona-2,4,6,8-tetraenoic acid 
with an additional carbon C16. To make a long story short this ligand was 
originally deposited missing this extra methyl goup in 1999 (as part of 3CBS) 
and then apparently updated in 2011 by the PDB. 

 
(the relevant lines in the cif are 
snip 
R12 C16 C16 C 0 1 N N N ?      ?      ?      -6.631 1.502  0.990  C16 R12 44  
R12 H1  H1  H 0 1 N N N ?      ?      ?      -6.602 1.511  2.080  H1  R12 45  
R12 H23 H23 H 0 1 N N N ?      ?      ?      -6.422 2.503  0.613  H23 R12 46  
R12 H24 H24 H 0 1 N N N ?      ?      ?      -7.619 1.186  0.656  H24 R12 47  
snip  

 
with the ? ? ? indicating that refined coordinates were not available at the 
time of the update. There was initially an explanation line at the end of the 
cif: 

 
snip 
R12 Other modification 2011-10-25 RCSB CS 'add missing methyl group, 
re-define bond order based on publication' 
snip 

 
But this has mutated for some reason (premature stop codon?) over the past 
year to the following. 

 
snip   
R12 Other modification 2011-10-25 RCSB  
snip 

 
Obviously the full correct ligand could not have been incorporated into the 
PDB entry coordinates without these undergoing a full obsolete - supersede 
process (somewhat embarrassing perhaps as one author is now a wwPDB PI ;) 

 
But it is frustrating for users of the PDB that in such cases easily 
correctable

Re: [ccp4bb] Comparison of Water Positions across PDBs

[MARTYN SYMMONS]

Thank you for that, Rachel

Even though the tone of your comment does not suggest that you want to carry on 
a dialogue about this, I thought I would reply in any case - since dialogue is 
what this forum is supposed to be about.

Thing is,  I was sort of looking for an explanation of why the rule was adopted 
that waters were to be renumbered from N to C terminus. If this is not 
functioning to put the waters in register across a set of related structures 
then it seems somewhat arbitrary. And other schemes might be suggested to be 
better for the usability and interpretation of the structural data. 

In some cases there are only a few waters but in many structures the wwPDB 
partners renumber hundreds. And this process makes it difficult for authors to 
check the final deposited structure against the output of their refinement. 

I have to say that I agree with other contributors to this thread. It would be 
much better to let the refinement program authors agree on a default water 
numbering scheme. And then maintain that through deposition. 

I thought of six possible schemes before breakfast... one of my favourites was 
to order by B-factor - which might appeal to crystallographers. Another was to 
give priority to those in the coordination sphere of any metal ions - these 
actually get priority in the PDB as they are included in the LINKS records 
above the coordinates. These coordinated waters are often refined together with 
the metals and so it would make sense to move them closer to their friendly 
ion.    

And of course one other clearly suitable option would be to leave the waters in 
the authors' preferred order - chosen with help from their refinement suite. 
This is what happens during deposition with the residues of the polymers - 
(provided the authors chainids are suitably chosen). Following your link the 
rule for polymers is that: 'If the coordinate residue numbers, as provided by 
the author, are unique and sequential within a particular chain ID, the 
residues will not be renumbered.' 

I'm presuming that if the authors have a preferred suitable set of water 
numbers then that would be maintained similarly?

Perhaps that is what is happening in the cases I notice that do not follow 
wwPDB rules?

On Friday I was looking at TIRAP structures and in 3ub2 the protein construct 
starts at residue 78 and its final residue is 221 - but the associated DTT is 
labelled back at residue 1 in the same chain. Then the first ten out of eleven 
waters are residues 2 to 11 but then oddly the eleventh water is residue 222. 
Is there a difference in this C-terminal water compared with the N-terminal 
ones? I imagined it was perhaps maintained to fit in with the associated 
publication - or maybe started out life modelled as a metal ion - unfortunately 
I can find no mention of it in the paper. 

But, regardless of this distracting feature,  surely this entry does not 
conform to the expected numbering scheme you mentioned as the wwPDB standard: 
polymer - heterogen - water? 

Yours perplexedly
 Martyn 



From: Rachel Kramer Green kra...@rcsb.rutgers.edu
To: CCP4BB@JISCMAIL.AC.UK 
Sent: Friday, 1 November 2013, 20:18
Subject: Re: [ccp4bb] Comparison of Water Positions across PDBs
 


In PDB format files, each polymer is assigned a unique chain ID. Chain IDs for 
all bound moieties and waters are assigned based on their proximity (number of 
contacts) to the nearest polymer. Once the polymers and non-polymer residues 
associated with them are assigned chain IDs, they are also assigned unique 
residue numbering with the order polymer residues, ligands and then waters. 

Please see: http://www.wwpdb.org/procedure.html#toc_4

The wwPDB has established this rule to improve the usability and
  interpretation of the structural data. Assigning the same chain ID
  for all moieties associated with a polymer enables rapid and
  uniform identification of feature analysis.

Sincerely,
Rachel Green

 


 
Rachel Kramer Green, Ph.D.
RCSB PDB
kra...@rcsb.rutgers.edu
 
 
Twitter: https://twitter.com/#!/buildmodels
Facebook: http://www.facebook.com/RCSBPDB
 
On 10/30/2013 8:09 AM, Eugene Krissinel wrote:

This is to be answered by PDB people, who definitely read BB :) Would be nice 
to have a tool common between CCP4/Phenix and the PDB which sorts this out 
Eugene On 30 Oct 2013, at 12:09, Andreas Förster wrote: 
Dear all, this water discussion is flowing increasingly towards a place where 
I feel a bit out of my depth. What is the convention for numbering water 
molecules?  Is there preference for: - putting waters into a separate chain (W 
for water or S for solvent)?
- splitting waters according to the peptide chains in the structure?
- appending all waters to chain A? Thanks. Andreas On 30/10/2013 11:57, MARTYN 
SYMMONS wrote: 
At deposition the PDB runs a script that renumbers authors'  waters
according to a scheme based on the residue

Re: [ccp4bb] Comparison of Water Positions across PDBs

[MARTYN SYMMONS]

Thanks Bernhard 
  you have helpfully distinguished between the two processes - there is 
certainly a movement of waters to symmetry replacements closer to a chain - and 
that gets documented in Remark 525 of the PDB file returned to authors - 
although then it is stripped out, I think, before the entry is released.

 But generally a renumbering is applied to all the waters - these are not 
actually moved but are re-ordered. And of course the number count of them may 
change - in order to accommodate any waters that are swapped in or out of the 
chain during the symmetry operation.

 Currently I don't think authors are given access to an audit of what is 
happening - they can of course check their favourite waters by a 
superimposition. Still have to say best way to avoid errors would be to check 
symmetry and re-order at the end of refinement, pre-deposition. 

All the best 
  Martyn 



 From: Bernhard Rupp hofkristall...@gmail.com
To: CCP4BB@JISCMAIL.AC.UK 
Sent: Monday, 4 November 2013, 14:50
Subject: Re: [ccp4bb] Comparison of Water Positions across PDBs
 


As far as confusion as a result of PDB renumbering is concerned: It was useful 
to run the old REM525 standalone program (I think I got it from PDBe/Kim 
Hendrick) at the end of solvent building. It does what the PDB did with water 
renumbering when creating the REMARK 525 (probably based on ccp4 contact with 
additions). Is there an updated standalone PDB tool available one can use these 
days to avoid at least that issue? 
 
Thx, BR
 
From:CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of MARTYN 
SYMMONS
Sent: Montag, 4. November 2013 15:17
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] Comparison of Water Positions across PDBs
 
Thank you for that, Rachel
 
Even though the tone of your comment does not suggest that you want to carry on 
a dialogue about this, I thought I would reply in any case - since dialogue is 
what this forum is supposed to be about.
 
Thing is,  I was sort of looking for an explanation of why the rule was adopted 
that waters were to be renumbered from N to C terminus. If this is not 
functioning to put the waters in register across a set of related structures 
then it seems somewhat arbitrary. And other schemes might be suggested to be 
better for the usability and interpretation of the structural data. 
 
In some cases there are only a few waters but in many structures the wwPDB 
partners renumber hundreds. And this process makes it difficult for authors to 
check the final deposited structure against the output of their refinement. 
 
I have to say that I agree with other contributors to this thread. It would be 
much better to let the refinement program authors agree on a default water 
numbering scheme. And then maintain that through deposition. 
 
I thought of six possible schemes before breakfast... one of my favourites was 
to order by B-factor - which might appeal to crystallographers. Another was to 
give priority to those in the coordination sphere of any metal ions - these 
actually get priority in the PDB as they are included in the LINKS records 
above the coordinates. These coordinated waters are often refined together with 
the metals and so it would make sense to move them closer to their friendly 
ion.    
 
And of course one other clearly suitable option would be to leave the waters in 
the authors' preferred order - chosen with help from their refinement suite. 
This is what happens during deposition with the residues of the polymers - 
(provided the authors chainids are suitably chosen). Following your link the 
rule for polymers is that: 'If the coordinate residue numbers, as provided by 
the author, are unique and sequential within a particular chain ID, the 
residues will not be renumbered.' 
 
I'm presuming that if the authors have a preferred suitable set of water 
numbers then that would be maintained similarly?
 
Perhaps that is what is happening in the cases I notice that do not follow 
wwPDB rules?
 
On Friday I was looking at TIRAP structures and in 3ub2 the protein construct 
starts at residue 78 and its final residue is 221 - but the associated DTT is 
labelled back at residue 1 in the same chain. Then the first ten out of eleven 
waters are residues 2 to 11 but then oddly the eleventh water is residue 222. 
Is there a difference in this C-terminal water compared with the N-terminal 
ones? I imagined it was perhaps maintained to fit in with the associated 
publication - or maybe started out life modelled as a metal ion - unfortunately 
I can find no mention of it in the paper. 
 
But, regardless of this distracting feature,  surely this entry does not 
conform to the expected numbering scheme you mentioned as the wwPDB standard: 
polymer - heterogen - water? 
 
Yours perplexedly
 Martyn 




From:Rachel Kramer Green kra...@rcsb.rutgers.edu
To: CCP4BB@JISCMAIL.AC.UK 
Sent: Friday, 1 November 2013, 20:18
Subject

Re: [ccp4bb] Comparison of Water Positions across PDBs

[MARTYN SYMMONS]

At deposition the PDB runs a script that renumbers authors'  waters according 
to a scheme based on the residue they are nearest from N to C terminus along 
each chain. This renumbering started  when waters were assigned to 
macromolecular chains rather than getting a chain id of their own.  I have 
failed to find the rationale explained in any PDB documents - but it could be 
motivated by this sort of consideration when waters from different chains or 
entries are to be compared. Having said that I do not know if there are any 
cases where this approach has successfully matched waters. ..


However an associated step which is certainly a help is that, in the case of 
multiple chains, the crystal symmetry is applied to replace waters with their 
symmetry equivalent position if it is closer to a different chain.

I believe a freely available program implementing a similar approach is 
WATERTIDY in CCP4 which might be a good place to start.  It gives a pretty 
complete output, detailing residues actually H-bonded to the waters, and you 
could parse that for further analysis and comparisons.

Best wishes.
  Martyn   


 From: Joel Sussman joel.suss...@weizmann.ac.il
To: CCP4BB@JISCMAIL.AC.UK 
Sent: Wednesday, 30 October 2013, 6:49
Subject: Re: [ccp4bb] Comparison of Water Positions across PDBs
 


For detailed examination of this topic, see: 


Koellner, G., Kryger, G., Millard, C. B., Silman, I., Sussman, J. L.  Steiner, 
T. (2000). 
Active-site gorge and buried water molecules in crystal structures of 
acetylcholinesterase from Torpedo californica. Journal of Molecular Biology 
296, 713-735.

http://www.ncbi.nlm.nih.gov/pubmed/10669619

best regards,
Joel

On 30 Oct 2013, at 01:35, Ed Pozharski epozh...@umaryland.edu wrote:

http://www.ccp4.ac.uk/html/watertidy.html


On 10/29/2013 04:43 PM, Elise B wrote:

Hello,

I am working on a project with several (separate) structures of the same 
protein. I would like to be able to compare the solvent molecules between the 
structures, and it would be best if the waters that exist in roughly the same 
position in each PDB share the
 same residue number. Basically, I want to compare solvent molecule coordinates 
and assign similar locations the same name in each structure.

What would be the best strategy for re-numbering the water molecules such 
that those with similar coordinates in all the structures receive the same 
residue number? I'd appreciate any suggestions.

Elise Blankenship



-- 
Oh, suddenly throwing a giraffe into a volcano to make water is crazy?
   Julian, King of Lemurs

Re: [ccp4bb] Problematic PDBs

[MARTYN SYMMONS]

As a postscript it might be worth mentioning one problematic ligand that 
suggested to me a way to correct some of the errors mentioned in this thread
 
R12 is indicated as 9-(4-HYDROXY-2,6-DIMETHYL-PHENYL)-3 in the  most recent 
Coot monomer library. But in the PDB ligand description it is 
9-(4-hydroxy-2,3,6-trimethylphenyl)-3,7-dimethylnona-2,4,6,8-tetraenoic acid 
with an additional carbon C16. To make a long story short this ligand was 
originally deposited missing this extra methyl goup in 1999 (as part of 3CBS) 
and then apparently updated in 2011 by the PDB.

(the relevant lines in the cif are
snip
R12 C16 C16 C 0 1 N N N ?      ?      ?      -6.631 1.502  0.990  C16 R12 44 
R12 H1  H1  H 0 1 N N N ?      ?      ?      -6.602 1.511  2.080  H1  R12 45 
R12 H23 H23 H 0 1 N N N ?      ?      ?      -6.422 2.503  0.613  H23 R12 46 
R12 H24 H24 H 0 1 N N N ?      ?      ?      -7.619 1.186  0.656  H24 R12 47 
snip 

with the ? ? ? indicating that refined coordinates were not available at the 
time of the update. There was initially an explanation line at the end of the 
cif:

snip
R12 Other modification 2011-10-25 RCSB CS 'add missing methyl group, 
re-define bond order based on publication'
snip

But this has mutated for some reason (premature stop codon?) over the past year 
to the following.

snip  
R12 Other modification 2011-10-25 RCSB 
snip

Obviously the full correct ligand could not have been incorporated into the PDB 
entry coordinates without these undergoing a full obsolete - supersede process 
(somewhat embarrassing perhaps as one author is now a wwPDB PI ;)

But it is frustrating for users of the PDB that in such cases easily 
correctable errors are not actually updated by the authors. Would it not be 
helpful if there were a mechanism to make and track useful improvements in 
deposited structures? - Perhaps suggested by members of the community to the 
authors. 

These changes could be considered as 'corrigenda' and could be documented and 
tracked - complete with an explanation of the reasoning behind the change and 
attributing the motivation and origin of the improvement.

This would be a good way for the wider scientific community (who maybe do not 
read this bulletin board) to access the best current model without the authors 
suffering the full process of retracting and redepositing their PDB entry. The 
test for obsoleting would then be the same as for a paper - that the change 
invalidates a fundamental interpretation of the data. 

All the best
  Martyn 



 From: Pavel Afonine pafon...@gmail.com
To: CCP4BB@JISCMAIL.AC.UK 
Sent: Sunday, 20 October 2013, 19:49
Subject: Re: [ccp4bb] Problematic PDBs
 


Hello,

just for the sake of completeness: this paper lists a bunch of known 
pathologies (I would not be surprised if they've been remediated by now):

http://www.phenix-online.org/papers/he5476_reprint.pdf


Pavel



On Thu, Oct 17, 2013 at 6:51 AM, Lucas lucasbleic...@gmail.com wrote:

Dear all,

I've been lecturing in a structural bioinformatics course where graduate 
students (always consisting of people without crystallography background to 
that point) are expected to understand the basics on how x-ray structures are 
obtained, so that they know what they are using in their bioinformatics 
projects. Practices include letting them manually build a segment from an 
excellent map and also using Coot to check problems in not so good structures.

I wonder if there's a list of problematic structures somewhere that I could 
use for that practice? Apart from a few ones I'm aware of because of (bad) 
publicity, what I usually do is an advanced search on PDB for entries with 
poor resolution and bound ligands, then checking then manually, hopefully 
finding some examples of creative map interpretation. But it would be nice to 
have specific examples for each thing that can go wrong in a PDB construction.

Best regards,
Lucas

Re: [ccp4bb] Docking models into low-res SAD map

[Martyn Symmons]

Hi Oliver
  a few years ago I used the ccp4 FFFEAR program from Kevin Cowtan for 
this sort of domain search. It was very good but also quite 
labour-intensive. I have not followed up since to see if it was updated 
or the functionality was included in Buccaneer.


cheers
Martyn


--
Martyn F. Symmons
Dept Veterinary Medicine
Cambridge

On 18/09/2013 23:48, Oliver Clarke wrote:

Hi all,

Can anyone recommend software to dock previously solved domains into a (very) 
low-resolution experimentally phased map?

I am working on a rather marginal case where this would be useful - very large 
assembly (500kDA per ASU), native goes to 6.9A, and the best derivative goes to 
8A with SAD phases from a heavy atom cluster to 11. Unfortunately no NCS is 
present.

The map is (for the resolution!) not too bad - the solvent boundary is clear 
after SHELXE, and I can manually place a previously solved structure and get a 
reasonably convincing fit (and the fit I obtain agrees with that previously 
determined using CryoEM by another group).

There are a couple of other domains that I would like to place, and I believe I 
have some idea of where they are, but manual fitting is somewhat subjective, so 
I'd like an automated routine for doing the same - something like the 
functionality that ADP_EM provides when working with CryoEM maps. Does 
something like this exist? I tried using ADP_EM but it gives a segfault when 
used with a crystallographic density map.

I have tried using MOLREP to look for the model in the map to no avail, and 
PHASER didn't work either.

On another note, if anyone has any tips for density modification/phase 
extension in this resolution range I would love to hear them - haven't had a 
whole lot of luck with PARROT, DM, or SOLOMON, the maps seem stuck at ~10A 
despite data going to 8. I tried using SHARP with the SPHCLUSTER tag on, but it 
gave no improvement over what I obtain treating the cluster as a super atom in 
SHELXD.

Many thanks in advance!

Oliver.

Re: [ccp4bb] What kind of reflection data to deposit to PDB

[Martyn Symmons]

Sorry to contradict,

But the mmCIF data model certainly does not seem to require hkl unique 
within the reflection data.


Like CIF the mmCIF formalism has been developed to allow a complete 
description of a diffraction experiment and the data arising from it. 
There is a full description at 
http://mmcif.pdb.org/dictionaries/mmcif_pdbx_v40.dic/Categories/diffrn_refln.html 
(I am grateful to Rachel Kramer Green at the RCSB for pointing out these 
links to the dictionary and the papers describing its development).


Having chosen mmCIF for the archive and then not using its flexibility 
seems a bit like having your cake and NOT eating it.


It is strange to hear on a discussion board that recently considered the 
advantages of depositing complete image data, that a case will have to 
be made for allowing the deposition of full unmerged datasets.


++Martyn


On 16/09/2013 14:03, Gerard DVD Kleywegt wrote:

Dear all,

At present, unmerged data cannot be handled properly as part of a PDB 
deposition. One reason for this is that changes to the mmCIF/PDBx data 
model will be required (at the moment, hkl must be unique within the 
reflection data, which is logical for merged data but precludes 
handling of unmerged data). There are other (easier to resolve) issues 
to work out, e.g. having to do with file naming and distribution via 
the wwPDB ftp archive.


The wwPDB partners are presently focusing all efforts on rolling out 
the new joint deposition and annotation system. Once this system is 
reasonably stable we will look into accepting/validating/distributing 
new kinds of data. This concerns not only unmerged Is for X-ray but 
also unassigned NOE peak lists for NMR. We will seek the advice of the 
corresponding wwPDB VTFs (Validation Task Forces) to help define the 
data items that need to be captured, how the data should be processed 
by wwPDB, and what kind(s) of validation is/are required.


--Gerard

---
Gerard J. Kleywegt, PDBe, EMBL-EBI, Hinxton, UK
ger...@ebi.ac.uk . pdbe.org
Secretary: Pauline Haslam pdbe_ad...@ebi.ac.uk




On Thu, 5 Sep 2013, Raji Edayathumangalam wrote:


Hi Folks,

Sorry for the non-ccp4 post.

I am trying to determine what is the best form of unmerged reflection 
data

to deposit to the PDB. I have single wavelength anomalous data for my
structure and I have two flavors of scaled files from the same exact 
set of
diffraction images: (1) data indexed and scaled in p1, and (2) data 
indexed
in p222, scaled in Scalepack using the no merge original index 
option and
converted to .mtz since the unit cell in the header of the output 
.sca file

was missing.

The space group for the dataset is p212121.

Please could you let me know what might be the best approach.

Many thanks and cheers,
Raji

--
Raji Edayathumangalam
Instructor in Neurology, Harvard Medical School
Research Associate, Brigham and Women's Hospital
Visiting Research Scholar, Brandeis University




Best wishes,

--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 !
**

Re: [ccp4bb] What kind of reflection data to deposit to PDB

[MARTYN SYMMONS]

Hi there,
To my knowledge the PDB does not include unmerged data in entries - you are 
required to deposit the data that the coordinates were refined against. 

I believe you can deposit other data which will be archived and can be 
requested by other users - problem is that none of the distribution web sites 
fesses up to the existence of these extra datasets as part of the entries - so 
users of your structure will be ignorant of them.

You could ask for an authors' remark to be included in the appropriate part of 
your coordinate header to alert users to the archived material. 

Currently you can upload data as mtz files and this format would of course be 
most that would be the most convenient. Of course the mmcif format could 
include the deposition of unmerged data.

Currently some distributed mmcif SF files in fact have multiple datasets - so 
for example relating to other crystals or other wavelengths. So you could 
simply provide your unmerged data as an mmcif block to include at the end of 
the dataset used for refinement. I'm guessing eds and pdbredo just pick up the 
first block as representative of the entry. But you can include other stuff 
after it. Again it would be useful to ask for explanatory remarks to be 
included for users of the data.

For example, the authors of the excellent structure 3ne5 submitted a 'phasing 
dataset' with their SFs. This appears (after 'Format standardization' 
2013-08-28) at the end of the native refinement dataset and I imagine refers to 
a SeMet crystal. (Although the values are so small I think they are perhaps the 
estimated intensities of just the anomalous-substructure? If you are 
interested, below is a bit from that file starting with the end of the native 
data - I imagine the negative sigmas indicate absent reflections).

You can check the mmcif dictionary at 
http://mmcif.rcsb.org/dictionaries/mmcif_pdbx.dic/Categories/refln.html

to see whether you can find an appropriate category for your data. I hope the 
PDB will be happy to spend the time required to negotiate with you an 
appropriate representation (although I was told that time constraints often 
mitigate against this).

all the best
   Martyn 


snip
1 1 1   47    1   25 o    306.1  133.1 1034.6586 1520.8234 
1 1 1   47    1   28 o    348.2  137.8 1795.0702 1421.0972 
#END
data_r3ne5Asf
# 
_cell.entry_id      3ne5 
_cell.length_a      160.076 
_cell.length_b      160.076 
_cell.length_c      680.069 
_cell.angle_alpha   90.000 
_cell.angle_beta    90.000 
_cell.angle_gamma   120.000 
# 
_diffrn.id                  1 
_diffrn.crystal_id          1 
_diffrn.ambient_temp        ? 
_diffrn.crystal_treatment   ? 
_diffrn.details             '  SeMet phasing set' 
# 
_diffrn_radiation_wavelength.id           1 
_diffrn_radiation_wavelength.wavelength   0.97920 
# 
_entry.id   3ne5 
# 
_exptl_crystal.id   1 
# 
_reflns_scale.group_code   1 
# 
_symmetry.entry_id               3ne5 
_symmetry.space_group_name_H-M   'H 3 2' 
# 
#
loop_
_refln.crystal_id
_refln.wavelength_id
_refln.scale_group_code
_refln.index_h
_refln.index_k
_refln.index_l
_refln.status
_refln.pdbx_I_plus
_refln.pdbx_I_plus_sigma
_refln.pdbx_I_minus
_refln.pdbx_I_minus_sigma
1 1 1    1    0  -29  o       13.6      1.1        0.0     -1.0 
1 1 1    1    0  -26  o        4.3      0.4        0.0     -1.0 
1 1 1    1    0  -23  o       13.1      0.8        0.0     -1.0 
1 1 1    1    0  -20  o      141.7      5.0        0.0     -1.0 
1 1 1    1    0  -17  o      305.0     10.6        0.0     -1.0 
1 1 1    2   -1   15  o      189.0      6.2      178.5      6.8
snip



 From: Raji Edayathumangalam r...@brandeis.edu
To: CCP4BB@JISCMAIL.AC.UK 
Sent: Friday, 6 September 2013, 3:42
Subject: [ccp4bb] What kind of reflection data to deposit to PDB
 


Hi Folks,

Sorry for the non-ccp4 post.

I am trying to determine what is the best form of unmerged reflection data to 
deposit to the PDB. I have single wavelength anomalous data for my structure 
and I have two flavors of scaled files from the same exact set of diffraction 
images: (1) data indexed and scaled in p1, and (2) data indexed in p222, scaled 
in Scalepack using the no merge original index option and converted to .mtz 
since the unit cell in the header of the output .sca file was missing. 

The space group for the dataset is p212121. 

Please could you let me know what might be the best approach.

Many thanks and cheers,
Raji

-- 
Raji Edayathumangalam
Instructor in Neurology, Harvard Medical School
Research Associate, Brigham and Women's Hospital
Visiting Research Scholar, Brandeis University

Re: [ccp4bb] Fractional coordinate shift with two-character chain names?

[MARTYN SYMMONS]

Hi Martyn 
   can you give me a reference where the rationale for this preference for 
mmcif is set out? I tried a pubmed search but cannot find anything. 
  thanks
   regards
   Martyn 



 From: Martyn Winn martyn.w...@stfc.ac.uk
To: CCP4BB@JISCMAIL.AC.UK 
Sent: Monday, 2 September 2013, 10:24
Subject: Re: [ccp4bb] Fractional coordinate shift with two-character chain 
names?
 

Hi,

There have been many discussions along these lines over the years.  The problem 
is understood well. The favoured solution is the use of mmCIF (as posted 
several times on the BB). But I would rather leave it to those officially 
involved to comment on the pros and cons. On the practical side, pdbcur will 
work equally well with mmCIF files.

Cheers
Martyn

*
*   Dr. Martyn Winn
*   STFC Daresbury Laboratory, Daresbury, Warrington, WA4 4AD, U.K.
*   Tel: +44 1925 603455 (DL)   or   +44 1235 567865 (RcaH)
*   E-mail: martyn.w...@stfc.ac.ukmailto:martyn.w...@stfc.ac.uk       Skype: 
martyn.winn
*


From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Dirk 
Kostrewa
Sent: 02 September 2013 09:38
To: ccp4bb
Subject: Re: [ccp4bb] Fractional coordinate shift with two-character chain 
names?

Dear Martyn, Pavel and other interested,

I think, an official extension by the PDB to two characters for the chain names 
and 5 digits for residues would really help. I'm currently working on a 
structure with 6x15 chains (through NCS) - it is huge and only a few programs 
can handle this by extending the PDB format.
The last PDB format revision 
3.3http://www.wwpdb.org/documentation/format33/sect9.html#ATOM still only 
allows one character for the chain name and four digits for the residue number.
More bigger structures will be published in the future and an official 
human-readable extended PDB format would really help.

Cheers,

Dirk.

Am 30.08.13 18:14, schrieb MARTYN SYMMONS:
Hold your horsemen!
Does not this option save us from 'formatagedon'?
We currently only have single letters or numbers for chains. But we could 
easily agree to switch to double letters. And long chains can be a sequence of 
letter number permutations such as A1, A2, A3 etc (actually I notice single 
numbers are allowed for the PDB - although are deprecated until all the letters 
have been used).
We could allow the first character to be a number as well - so 11 12 13 as a 
valid sequence.for a single polymer.
Conversely we could expand the atom identifier to include letters as is the 
case with most computing identifiers - however not many programs seem to pay 
attention to the atom 'numbers' in any case.

Cheers
  Martyn

Martyn Symmons (not Winn)
Cambridge


From: Dirk Kostrewa 
kostr...@genzentrum.lmu.demailto:kostr...@genzentrum.lmu.de
To: CCP4BB@JISCMAIL.AC.UKmailto:CCP4BB@JISCMAIL.AC.UK
Sent: Friday, 30 August 2013, 15:36
Subject: Re: [ccp4bb] Fractional coordinate shift with two-character chain 
names?

Hi Martyn,

excellent - this worked!

Many thanks!

Cheers,

Dirk.

Am 30.08.13 16:04, schrieb Martyn Winn:
 IIRC the CCP4 library (i.e. mmdb) can handle 2-character chain names. There 
 may be something specific in pdbset which interferes. You can try pdbcur as 
 an alternative. Something like:

 pdbcur xyzin toxd_AA.pdb xyzout toxd_out.pdb eof
 translate * frac 0 0.2 0
 end
 eof

 I just tried it on a little example, and it works for me.

 Cheers
 Martyn

 -Original Message-
 From: CCP4 bulletin board 
 [mailto:CCP4BB@JISCMAIL.AC.UKmailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of
 Dirk Kostrewa
 Sent: 30 August 2013 14:41
 To: ccp4bb
 Subject: [ccp4bb] Fractional coordinate shift with two-character chain
 names?

 Dear CCP4ers,

 I want to apply a fractional coordinate shift along a polar b-axis with
 coordinates that have non-standard two-character chain names, such as
 AA, AB, and so forth. Unfortunately, neither the old USF moleman2
 nor the actual CCP4 pdbset can handle these chain names. To my
 knowledge, only COOT and PHENIX can cope with them.
 Before I start writing my own little jiffy, is there a quick way to use
 COOT or PHENIX to apply a fractional coordinate shift, or could you
 tell me, which other program I can use in this special case?

 Best regards,

 Dirk.

 --

 ***
 Dirk Kostrewa
 Gene Center Munich
 Department of Biochemistry
 Ludwig-Maximilians-Universität München
 Feodor-Lynen-Str. 25
 D-81377 Munich
 Germany
 Phone:     +49-89-2180-76845
 Fax:     +49-89-2180-76999
 E-mail:    kostr...@genzentrum.lmu.demailto:kostr...@genzentrum.lmu.de
 WWW:    www.genzentrum.lmu.dehttp://www.genzentrum.lmu.de
 ***

--

***
Dirk Kostrewa
Gene Center Munich
Department of Biochemistry
Ludwig-Maximilians-Universität München
Feodor-Lynen-Str. 25
D-81377 Munich
Germany
Phone:     +49-89-2180-76845
Fax:     +49-89-2180

Re: [ccp4bb] Fractional coordinate shift with two-character chain names?

[MARTYN SYMMONS]

Hold your horsemen!
Does not this option save us from 'formatagedon'?
We currently only have single letters or numbers for chains. But we could 
easily agree to switch to double letters. And long chains can be a sequence of 
letter number permutations such as A1, A2, A3 etc (actually I notice single 
numbers are allowed for the PDB - although are deprecated until all the letters 
have been used). 
We could allow the first character to be a number as well - so 11 12 13 as a 
valid sequence.for a single polymer.
Conversely we could expand the atom identifier to include letters as is the 
case with most computing identifiers - however not many programs seem to pay 
attention to the atom 'numbers' in any case. 

Cheers

  Martyn 

Martyn Symmons (not Winn)
Cambridge



 From: Dirk Kostrewa kostr...@genzentrum.lmu.de
To: CCP4BB@JISCMAIL.AC.UK 
Sent: Friday, 30 August 2013, 15:36
Subject: Re: [ccp4bb] Fractional coordinate shift with two-character chain 
names?
 

Hi Martyn,

excellent - this worked!

Many thanks!

Cheers,

Dirk.

Am 30.08.13 16:04, schrieb Martyn Winn:
 IIRC the CCP4 library (i.e. mmdb) can handle 2-character chain names. There 
 may be something specific in pdbset which interferes. You can try pdbcur as 
 an alternative. Something like:

 pdbcur xyzin toxd_AA.pdb xyzout toxd_out.pdb eof
 translate * frac 0 0.2 0
 end
 eof

 I just tried it on a little example, and it works for me.

 Cheers
 Martyn

 -Original Message-
 From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of
 Dirk Kostrewa
 Sent: 30 August 2013 14:41
 To: ccp4bb
 Subject: [ccp4bb] Fractional coordinate shift with two-character chain
 names?

 Dear CCP4ers,

 I want to apply a fractional coordinate shift along a polar b-axis with
 coordinates that have non-standard two-character chain names, such as
 AA, AB, and so forth. Unfortunately, neither the old USF moleman2
 nor the actual CCP4 pdbset can handle these chain names. To my
 knowledge, only COOT and PHENIX can cope with them.
 Before I start writing my own little jiffy, is there a quick way to use
 COOT or PHENIX to apply a fractional coordinate shift, or could you
 tell me, which other program I can use in this special case?

 Best regards,

 Dirk.

 --

 ***
 Dirk Kostrewa
 Gene Center Munich
 Department of Biochemistry
 Ludwig-Maximilians-Universität München
 Feodor-Lynen-Str. 25
 D-81377 Munich
 Germany
 Phone:     +49-89-2180-76845
 Fax:     +49-89-2180-76999
 E-mail:    kostr...@genzentrum.lmu.de
 WWW:    www.genzentrum.lmu.de
 ***

-- 

***
Dirk Kostrewa
Gene Center Munich
Department of Biochemistry
Ludwig-Maximilians-Universität München
Feodor-Lynen-Str. 25
D-81377 Munich
Germany
Phone:     +49-89-2180-76845
Fax:     +49-89-2180-76999
E-mail:    kostr...@genzentrum.lmu.de
WWW:    www.genzentrum.lmu.de
***

Re: [ccp4bb] modified amino acids in the PDB

[MARTYN SYMMONS]

Hi Clemens
   I guess the reason you say 'arbitrary' is because there is no explanation of 
this rule decision? 
  It would be nice if some rationalization was available alongside the values 
given. So a sentence along the lines of 'we set the number owing to the 
following considerations' ? 

  However a further layer of variation is that the rule does not seem to be 
consistently applied
 - just browsing CYS modifications:
   iodoacetamide treatment gives a CYS with only 4 additional atoms but it is 
split off as  ACM.
   However some ligands much larger than 10 residues have been kept with the 
cysteine ( for example CY7 in 2jiv and NPH in 1a18.  
   My betting is that it depends on whether something has been seen 'going 
solo' as a non-covalent ligand previously so that it pops up as an atomic 
structural match with a pre-defined three-letter code.
  This would explain for example the ACM case which you might expect to occur 
in a modified Cys.  But it has also been observed as a non-polymer ligand in 
its own right so goes on as a separate modification?
   However to be honest I am not sure I have ever seen the rationale for this 
written down. 

  'Non-polymer' heterogens can turn up either linked or not. Once they are in 
the residues they have to make a call on which kind of backbone they will 
feature in within the pdb.
  That is why there is  'D5M' for non-polymer deoxyAMP. Also known as ' DA' 
when it is 'DNA-linking' but so far not fessing up to life under a third code 
as 'RNA-linking' 

Now is perhaps the time to ask for explanations of these nomenclature features 
before they become hard-wired in the new pdb deposition system (however there 
may be time - I refer you to my previous posting ;). 
  
 Cheers
    Martyn 
  
Dr Martyn Symmons
Cambridge



 From: Michael Weyand michael.wey...@uni-bayreuth.de
To: CCP4BB@JISCMAIL.AC.UK 
Sent: Monday, 8 July 2013, 10:03
Subject: [ccp4bb] modified amino acids in the PDB
 

Dear colleagues,

We deposited protein structures with modified lysine side chains and
were surprised that the PDB treats the modification as an independent
molecule, with a “LINK” record indicating the covalent bond – instead of
defining a modified residue (that’s what we had uploaded to the PDB).
Apparently, anything attached to an amino acid is considered an
independent molecule (and the lysine just called a regular lysine) if it
comprises more than 10 atoms (see below for the PDB guidelines).

I think that’s kind of arbitrary and would give all modified residue
also modified names – i.e. individual names for all modified lysines, as
it is done for acetyl- or methyl-lysines, for example. I wonder what
other people’s opinion is?!

Best regards

Clemens



This is in accordance to the wwPDB annotation guidelines
(http://www.wwpdb.org/procedure.html#toc_2).
*Modified amino acids and nucleotides* If an amino acid or nucleotide
is modified by a chemical group greater than 10 atoms, the residue will
be split into two groups: the amino acid/nucleotide group and the
modification. A link record will be generated between the amino
acid/nucleotide group and the modification. For modified amino acids and
nucleotides that were not split will follow standard atom nomenclature.

Re: [ccp4bb] modified amino acids in the PDB

[MARTYN SYMMONS]

Hiya Mark
   once again I find myself asking why not give the authors deposited file 
alongside any other 'cleaned-up' PDB files. 
 The authors' one could be foo.pdb_0 then after PDB heterogen annotation you 
get foo.pdb_1 - if the heterogens are subsequently handled differently then you 
could have foo.pdb_2 for the 'remediated' file. 
  This latter has certainly happened as there appear to be 'orphan' heterogen 
definitions in the pdb that are not currently used by any entries - most likely 
these were 'split' when atoms were taken out and associated with polymer 
entities, or in some cases, I notice, 'lumped' when they have been included 
with residues alongside to give a new larger heterogen.
  PDB file versioning would also give transparency in cases such as occupancy 
remediation when the pdb altered all occupancies that summed to 1.0 to enforce 
a  total of 1.0 (giving holes in the authors' density presumably) which may 
mystify the sharp-eyed user.    
 Currently there are the REV_DAT lines in the header but these give only the 
titles of changed cards not any detailed explanation. Likewise REVDAT only 
starts at foo.pdb_1 in my example - still missing the authors original intent.  
  all the best
   Martyn 


 From: Mark J van Raaij mjvanra...@cnb.csic.es
To: CCP4BB@JISCMAIL.AC.UK 
Sent: Tuesday, 9 July 2013, 15:23
Subject: Re: [ccp4bb] modified amino acids in the PDB
 

- really the only complicated case would be where a group is covalently linked 
to more than one amino acid, wouldn't it? Any case where only one covalent link 
with an is present could (should?) be treated as a special amino acid, i.e. 
like selenomethionine.
- groups without any covalent links to the protein are better kept separate I 
would think (but I guess this is stating the obvious).

Mark J van Raaij
Lab 20B
Dpto de Estructura de Macromoleculas
Centro Nacional de Biotecnologia - CSIC
c/Darwin 3
E-28049 Madrid, Spain
tel. (+34) 91 585 4616
http://www.cnb.csic.es/~mjvanraaij





On 9 Jul 2013, at 12:49, Frances C. Bernstein wrote:

 In trying to formulate a suggested policy on het groups
 versus modified side chains one needs to think about the
 various cases that have arisen.
 
 Perhaps the earliest one I can think of is a heme group.
 One could view it as a very large decoration on a side
 chain but, as everyone knows, one heme group makes four
 links to residues.  In the early days of the PDB we decided
 that heme obviously had to be represented as a separate group.
 
 I would also point out that nobody would seriously suggest that
 selenomethionine should be represented as a methionine with a
 missing sulfur and a selenium het group bound to it.
 
 Unfortunately all the cases that fall between selenomethionine
 and heme are more difficult.  Perhaps the best that one must
 hope for is that whichever representation is chosen for a
 particular case, it be consistent across all entries.
 
                          Frances
 
 P.S. One can also have similar discussions about the representation
 of microheterogeneity and of sugar chains but we should leave those
 for another day.
 
 =
                 Bernstein + Sons
 *   *       Information Systems Consultants
     5 Brewster Lane, Bellport, NY 11713-2803
 *   * ***
  *            Frances C. Bernstein
  *   ***      f...@bernstein-plus-sons.com
 ***     *
  *   *** 1-631-286-1339    FAX: 1-631-286-1999
 =
 
 On Tue, 9 Jul 2013, MARTYN SYMMONS wrote:
 
 Hi Clemens
    I guess the reason you say 'arbitrary' is because there is no explanation 
of this
 rule decision? 
   It would be nice if some rationalization was available alongside the 
values given.
 So a sentence along the lines of 'we set the number owing to the following
 considerations' ? 
   However a further layer of variation is that the rule does not seem to be
 consistently applied
  - just browsing CYS modifications:
    iodoacetamide treatment gives a CYS with only 4 additional atoms but it 
is split
 off as  ACM.
    However some ligands much larger than 10 residues have been kept with the 
cysteine
 ( for example CY7 in 2jiv and NPH in 1a18.  
    My betting is that it depends on whether something has been seen 'going 
solo' as a
 non-covalent ligand previously so that it pops up as an atomic structural 
 match with
 a pre-defined three-letter code.
   This would explain for example the ACM case which you might expect to 
occur in a
 modified Cys.  But it has also been observed as a non-polymer ligand in its 
 own right
 so goes on as a separate modification?
    However to be honest I am not sure I have ever seen the rationale for 
this written
 down. 
   'Non-polymer' heterogens can turn up either linked or not. Once they are 
in the
 residues they have to make a call on which kind of backbone they will 
 feature in
 within the pdb.
   That is why there is  'D5M

Re: [ccp4bb] PDB secondary structure assignments

[MARTYN SYMMONS]

Authors can supply the secondary structure assignments to the PDB and these 
trump the PDBs own determinations and will appear in the authors' entry.

Remarks 650 (Helix) and 700 (Sheet) are then included which allow the 
documentation of the origins of the secondary structure definitions and any 
useful details from the authors on the determination methods used (presumably 
down to the trivial ' this looked prettier to me than the DSSP ones').

For the record you can check the pdb annotation manual 
(http://deposit.rcsb.org/depoinfo/download/dp.pdf) that seems to indicate the 
secondary structure is cooked up by the in-house 'maxit' program.

However there are some delightful sketch drawings (pages 92 to 93) showing how 
the annotators are supposed to work stuff out with a pencil-and-paper.

This may explain cases where the annotators have included material in the 
secondary structure lines which were presumably helpful comments that should've 
been trimmed out 

For example in  entry 3pcy:

HELIX    1   A ALA A   52  SER A   56  1ONLY TURN OF HELIX IN MOLCULE      5    
SHEET    1   I 4 GLY A  10  VAL A  15  0                                        
SHEET    2   I 4 ILE A   1  ALA A   7 -1  N  GLY A   6   O  ALA A  13           
SHEET    3   I 4 GLU A  25  ASN A  32  1  O  LYS A  26   N  ILE A   1           
SHEET    4   I 4 GLY A  67  LEU A  74 -1  O  GLU A  68   N  ASN A  31           

which is, I suppose useful, if you were looking for the 'only turn of helix in 
molcule'?

Of course we can expect these quirks to be a thing of the past when the long 
promised wwPDB 'Common Deposition and Annotation Tool' is rolled out. 

This Common Tool is apparently now 'nearing readiness for testing by external 
users' (http://www.wwpdb.org/wwpdbac-pages/wwPDBAC2012_report.pdf).

Hmm...
Previously advertised as:
'completion is on track for 2012.' 
(http://www.wwpdb.org/wwpdbac-pages/wwPDBAC_2011_Report.pdf)

or that 'substantial completion in 2011 is likely.' 
http://www.wwpdb.org/wwpdbac-pages/wwpdbac-2010-report.pdf


or perhaps more pertinently:

'In early 2010, the annotation module will be put into service,'

(http://www.wwpdb.org/wwpdbac-pages/wwpdbac-2010-report.pdf)


Perhaps as in many things it is often better to travel hopefully than to 
arrive... 

Cheers
  Martyn

Martyn Symmons
Cambridge UK



 From: Mark J van Raaij mjvanra...@cnb.csic.es
To: CCP4BB@JISCMAIL.AC.UK 
Sent: Wednesday, 3 July 2013, 16:00
Subject: Re: [ccp4bb] PDB secondary structure assignments
 

you are probably right, but the depositor can override the automatic secondary 
structure assignment by submitting his own. Perhaps this is what happened here.
I would recommend the original poster to generate them by hand by inspection of 
the structure plus density if possible - i.e. not trust any program...
Or, in case he wants to do it for a lot of structures and inspection of them 
all is not feasible, regenerate them all with the same version of his favourite 
program.


Mark J van Raaij
Lab 20B
Dpto de Estructura de Macromoleculas
Centro Nacional de Biotecnologia - CSIC
c/Darwin 3
E-28049 Madrid, Spain
tel. (+34) 91 585 4616
http://www.cnb.csic.es/~mjvanraaij





On 3 Jul 2013, at 16:46, Robbie Joosten wrote:

 Hi Miha,
 
 I thought the PDB actually uses DSSP. Perhaps it is a different version,
 there have been some new releases recently. Anyway, there is no reason why
 you should stick to the assignment of the PDB. If another program gives
 slightly different results you can use those as long as you make sure it is
 obvious which program you used (cite the program).  
 The next CCP4 release will have the official dssp (which is used to make the
 DSSP databank).
 
 Cheers,
 Robbie
 
 
 
 -Original Message-
 From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of
 Pavšic, Miha
 Sent: Wednesday, July 03, 2013 16:01
 To: CCP4BB@JISCMAIL.AC.UK
 Subject: [ccp4bb] PDB secondary structure assignments
 
 Dear CCP4BB members,
 
 what is the usual practice regarding secondary structure assignments when
 preparing publication figures of protein structures and topology diagrams
 from deposited PDB files? The deposited PDB files already contain such
 assignments in the header section (using PROMOTIF?). Should these
 assignments be obeyed or is it common to used other software/algorithms
 (e.g., DSSP and Stride). In my case assignments using DSSP result in
 slightly
 differ from PDB assignments in the regions of short alpha-helical
 structure
 (corresponding to stretch of 4 aa residues).
 
 Thank you for your suggestions.
 
 Regards,
 Miha
 
 
 **
 Miha Pavsic, Ph.D.
 University of Ljubljana
 Faculty of Chemistry and Chemical Technology Chair of Biochemistry Cesta v
 Mestni log 88a
 SI-1000 Ljubljana
 Slovenia
 
 e-mail miha.pav...@fkkt.uni-lj.si
 skype mihapavsic
 phone (lab) +386 1 2419 488
 fax +386 1 2419 487
 **

Re: [ccp4bb] showing electron density on an ipad

[MARTYN SYMMONS]

Dear Dave
    I have found the Molsoft ICM browser useful for showing students 
structures. There is a freely distributed version available for both ipad and 
android. I have not used it for maps but I believe it can display them. 
  SGC Oxford originally showed me this software and have great examples 
displaying their structures on their web site and in their PLoS articles. Sorry 
but  I have not used the ipad version - I think you have to pay a nominal fee 
for that.
 All the best
  Martyn    



 From: David Roberts drobe...@depauw.edu
To: CCP4BB@JISCMAIL.AC.UK 
Sent: Wednesday, 1 May 2013, 20:56
Subject: [ccp4bb] showing electron density on an ipad
 

Hello all,

So, I find an ipad is a wonderful device for teaching (any tablet really - but 
I'm partial to the ipad).  I can project it in a classroom, run pymol and a few 
other chemistry/biochemistry things, and really get the students interested in 
these subjects easily.  I actually don't have a laptop - and our classrooms are 
such that there are computers connected to the projectors but they have 
standard University software packages installed on them.

It would be very helpful if I could just display electron density using an 
ipad.  The pymol app will load a map (it is an option) - but when I take a map 
from my linux machines it doesn't work.  Any thoughts here?  Has anybody done 
this

Thanks

Dave

Re: [ccp4bb] Puzzling Structure

[MARTYN SYMMONS]

And as a footnote*
-
* I notice that, following discussion on this bulletin board, the space group 
of the 2wlv coordinate file has been updated.

 But at this point did it not occur to the PDB to change the space group in the 
structure factor file as well? So the coordinates indicate P21 21 2 but the 
structure factor file still says P21 21 21... Hmm consistency is highly 
over-rated...

Also the author's SF data listing starts with c axis reflections including 0, 
0, l odd as well as even, which is sort of a hint here



 From: MARTYN SYMMONS martainn_oshioma...@btinternet.com
To: CCP4BB@JISCMAIL.AC.UK 
Sent: Tuesday, 16 April 2013, 14:30
Subject: Re: [ccp4bb] Puzzling Structure
 


Just as a postscript.

It has been pointed out to me that the space group was correct in the uploaded 
PDB and in the mtz reflection file. And there was no editing of this data 
pre-deposition. 

I am sort of surprised how quickly people are to beat up on the authors in such 
cases since we do not have access to the full data and facts.

Having the uploaded data available would allow better checking. I'm mainly 
ignorant of the details, but isn't this the way sequence data is dealt with? - 
there are the unannotated sequence files which are gradually checked and 
assimilated (but not over-written).   

Cheers 
  Martyn 



 From: Michel Fodje michel.fo...@lightsource.ca
To: CCP4BB@JISCMAIL.AC.UK 
Sent: Monday, 15 April 2013, 17:19
Subject: Re: [ccp4bb] Puzzling Structure
 

Just to round up this topic, a bug report was submitted to PDBe concerning 
entry 2wlv and the PDB has how responded acknowledging the problem. An updated 
entry will be available in one week.

As pointed out by Savvas, it is very likely the CRYST1 record was manually 
edited prior to deposition resulting in the wrong spacegroup being parsed in by 
AutoDep and subsequent processing moving the waters around in the wrong space 
group. Since there is no logical reason for the authors to do this, it was 
probably inadvertent. I imagine somebody accidentally deleted a space between P 
21 21 2 and 18 and tried to fix
 it by adding it back, between 1 and 8.  

/Michel

-Original Message-
From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of
Edward A. Berry
Sent: April-14-13 9:59 AM
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] Puzzling Structure

Robbie Joosten wrote:
 Hi Martyn,

     I think the question is where the error was made - seeing the
 uploaded file would clear this up. But it seems unlikely to me that
 the depositor saw a huge R factor discrepancy at the end of refinement
and just blithely uploaded it.
 So scenario 3 :-
 PDB : we cannot reproduce your R factor with our programs Depositor
 :
 that's your problem mate - it was fine when it left me...up to you to sort
it...
     Which seems a sort of reasonable attitude to me.

 Not quite, the depositor has to give, i.e. type, the space group (example
depositions: https://www.ebi.ac.uk/pdbe- 
xdep/autodep/AutoDep?param=QovCsvhNv06Mpnr%2BvIkqqfuqeeBd8leFN
AVymZgS%2Fe7mULyfrCaTMN8jsyaGZUTyUDQyN3gMF3o%3D). Don't ask me
why, because it is clearly a source of error.


In my experience with RCSB autodep2, assuming the information was in the
uploaded pdb file, this information is already pre-filled and the depositor 
just
glances over to see it is correct. A missing or extra (1) might not be 
noticed.
So perhaps it is a parsing error, perhaps related to the different ways the
space group is represented on the CRYST1 card, and the different stringency
of different programs in interpreting the CRYST1 card.

But the validation report is included with the approval request letter, and
major discrepancies are noted in the text of the validation letter in case the
depositor is too busy to actually read the validation report.
So if the
 depositor read more than the first line or two of the letter he should
have known there was a problem.

Then there is the 2-week default release policy:

  No major issues were raised during data processing. A summary of some of
the key annotations
  in your entry is shown below. Please verify that these are correct. If we do
not hear from
  you within two weeks, we will consider this entry to have been approved by
you. The entry
  will then be released according to the release/hold instructions you have
provided.

If this 2-week default release is the rule even when there are issues, the
depositor may have put it aside to find the problem when time was available,
and waited too long and let the default release kick in.

eab

Re: [ccp4bb] Puzzling Structure

[MARTYN SYMMONS]

Just as a postscript.

It has been pointed out to me that the space group was correct in the uploaded 
PDB and in the mtz reflection file. And there was no editing of this data 
pre-deposition. 

I am sort of surprised how quickly people are to beat up on the authors in such 
cases since we do not have access to the full data and facts.

Having the uploaded data available would allow better checking. I'm mainly 
ignorant of the details, but isn't this the way sequence data is dealt with? - 
there are the unannotated sequence files which are gradually checked and 
assimilated (but not over-written).   

Cheers 
  Martyn 



 From: Michel Fodje michel.fo...@lightsource.ca
To: CCP4BB@JISCMAIL.AC.UK 
Sent: Monday, 15 April 2013, 17:19
Subject: Re: [ccp4bb] Puzzling Structure
 

Just to round up this topic, a bug report was submitted to PDBe concerning 
entry 2wlv and the PDB has how responded acknowledging the problem. An updated 
entry will be available in one week.

As pointed out by Savvas, it is very likely the CRYST1 record was manually 
edited prior to deposition resulting in the wrong spacegroup being parsed in by 
AutoDep and subsequent processing moving the waters around in the wrong space 
group. Since there is no logical reason for the authors to do this, it was 
probably inadvertent. I imagine somebody accidentally deleted a space between P 
21 21 2 and 18 and tried to fix it by adding it back, between 1 and 8.  

/Michel

-Original Message-
From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of
Edward A. Berry
Sent: April-14-13 9:59 AM
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] Puzzling Structure

Robbie Joosten wrote:
 Hi Martyn,

     I think the question is where the error was made - seeing the
 uploaded file would clear this up. But it seems unlikely to me that
 the depositor saw a huge R factor discrepancy at the end of refinement
and just blithely uploaded it.
 So scenario 3 :-
 PDB : we cannot reproduce your R factor with our programs Depositor :
 that's your problem mate - it was fine when it left me...up to you to sort
it...
     Which seems a sort of reasonable attitude to me.

 Not quite, the depositor has to give, i.e. type, the space group (example
depositions: https://www.ebi.ac.uk/pdbe- 
xdep/autodep/AutoDep?param=QovCsvhNv06Mpnr%2BvIkqqfuqeeBd8leFN
AVymZgS%2Fe7mULyfrCaTMN8jsyaGZUTyUDQyN3gMF3o%3D). Don't ask me
why, because it is clearly a source of error.


In my experience with RCSB autodep2, assuming the information was in the
uploaded pdb file, this information is already pre-filled and the depositor 
just
glances over to see it is correct. A missing or extra (1) might not be 
noticed.
So perhaps it is a parsing error, perhaps related to the different ways the
space group is represented on the CRYST1 card, and the different stringency
of different programs in interpreting the CRYST1 card.

But the validation report is included with the approval request letter, and
major discrepancies are noted in the text of the validation letter in case the
depositor is too busy to actually read the validation report.
So if the depositor read more than the first line or two of the letter he 
should
have known there was a problem.

Then there is the 2-week default release policy:

  No major issues were raised during data processing. A summary of some of
the key annotations
  in your entry is shown below. Please verify that these are correct. If we do
not hear from
  you within two weeks, we will consider this entry to have been approved by
you. The entry
  will then be released according to the release/hold instructions you have
provided.

If this 2-week default release is the rule even when there are issues, the
depositor may have put it aside to find the problem when time was available,
and waited too long and let the default release kick in.

eab

Re: [ccp4bb] Puzzling Structure

[MARTYN SYMMONS]

Dear Robbie

A shame then that these 'helpful' annotators did not make use of  Pavel's basic 
sanity on the space group (*mentioned below) and check back to the one listed 
in the uploaded PDB file. 

I often wonder why the PDB does not make the deposited coordinate file publicly 
available so that these sorts of issues can be checked and tracked. Depositors' 
coordinate files nowadays are the output of reputable, documented programs.  

The whole PDB data (excluding the EMDB that was recently merged into it) 
amounts to about a laptop hard drive's worth of data  - so surely space can be 
made for the deposited coordinates? (and restraint files which will be very 
useful for other workers including pdb-redo).

Structural genomic groups could also take a lead here by making their final 
refined coordinates and restraint files available alongside the PDB data. 

Having the depositors' uploaded data would help me understand other puzzling 
features of structures such as the current 4GRV.pdb which seems to have a list 
of TLS groups but contains not a single ANISOU line!...  

Cheers
  Martyn 

*In this particular case attempting to calculate R-factor using data and model 
files and making sure that the R you get is not twice as large as published one 
would entirely suffice -:)

Pavel



 From: Robbie Joosten robbie_joos...@hotmail.com
To: CCP4BB@JISCMAIL.AC.UK 
Sent: Friday, 12 April 2013, 22:57
Subject: Re: [ccp4bb] Puzzling Structure
 

Waters are moved during annotation using the perceived space group's
symmetry operation. So if the authors give the wrong space group, then the
annotation pipeline understandably messes things up. If the originally
uploaded PDB file was kept by PDBe, then the problem can be recovered quite
easily by the annotators. Perhaps the topic starter, Michel Fodje, can send
a bug report to PDBe. In my experience, the annotators are very helpful
resolving these matters.  

potential flame
Hoping that the depositors solve the problem by themselves, is probably in
vain: There are many crystallographers who do not read the CCP4BB (which is
a shame, really); they didn't notice the enormous amount of water related
bumps in their final model (which is in the validation report you get after
deposition and in REMARK 500 of the PDB file you have to approve); they also
didn't notice the huge number of symmetry-related bumps; the R-factors in
the PDB file are different from (and better than) the ones in Table 1. Also
notice that the paper was submitted on April 21st 2009 and the model was
deposited on June 29th 2009. Paper accepted on July 8th 2009. But I'm sure
the referees had a chance to properly assess the quality of the structure
model ;-)
/ potential flame

Cheers,
Robbie

P.S. It's pretty awesome that the problem was solved in less than 20 minutes
by the CCP4BB (that is, by Phoebe Rice)


 -Original Message-
 From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of
 Garib N Murshudov
 Sent: Friday, April 12, 2013 21:39
 To: CCP4BB@JISCMAIL.AC.UK
 Subject: Re: [ccp4bb] Puzzling Structure
 
 It is typo:
 R factor for p212121 - 0.4
                for p21212    - around 0.18
 
 Although water seem to have been moved around using p212121
 
 
 
 
 On 12 Apr 2013, at 16:33, Phoebe A. Rice wrote:
 
 
     Looks like a typo to me: if you change the CRYST space group record
 from P212121 to P21212, as the paper says it is, the packing problem goes
 away.
 
     ++
 
     Phoebe A. Rice
     Dept. of Biochemistry  Molecular Biology
     The University of Chicago
 
     773 834 1723; pr...@uchicago.edu
     http://bmb.bsd.uchicago.edu/Faculty_and_Research/
 
     http://www.rsc.org/shop/books/2008/9780854042722.asp
 
     
     From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] on behalf of
 Michel Fodje [michel.fo...@lightsource.ca]
     Sent: Friday, April 12, 2013 2:17 PM
     To: CCP4BB@JISCMAIL.AC.UK
     Subject: Re: [ccp4bb] Puzzling Structure
 
     By the way, you will need to show symmetry atoms to see the
 problem.
 
 
 
         -Original Message-
 
 
         From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK]
 On Behalf Of
 
 
         Michel Fodje
 
 
         Sent: April-12-13 1:14 PM
 
 
         To: CCP4BB@JISCMAIL.AC.UK
 
 
         Subject: [ccp4bb] Puzzling Structure
 
 
 
         Has anyone else noticed a problem with the structure  of the
 N-terminal
 
 
         capsid domain of HIV-2  PDB 2wlv.
 
 
 
         Load it up to in coot and navigate to residue B118.
 
 
 
 
 
         /Michel.
 
 
 Dr Garib N Murshudov
 Group Leader, MRC Laboratory of Molecular Biology Francis Crick Avenue
 Cambridge Biomedical Campus Cambridge
 CB2 0QH UK
 Email: ga...@mrc-lmb.cam.ac.uk
 Web http://www.mrc-lmb.cam.ac.uk http://www.mrc-lmb.cam.ac.uk/
 
 
 
 
 
 

Re: [ccp4bb] PDBe website update: What's new

[MARTYN SYMMONS]

Ah yes a new logo...

But it might be of interest to hear in this forum what has happened to the CCP4 
logo that used to appear on the PISA and Fold pages in acknowledgement of the 
support from the BBSRC on the underlying algorithms for these services.

It is only by acknowledgement of the contribution of projects such as CCP4 that 
we can establish a continuing funding stream for the improvement of scientific 
software. I distinctly remember that this was acknowledged by the presence of 
the CCP4 logo on appropriate pages in the old PDBe website. Surely there is 
still space for that acknowledgement too?

Yours perplexedly,

Martyn 

Dr Martyn Symmons
Cambridge



 From: Gary Battle bat...@ebi.ac.uk
To: CCP4BB@JISCMAIL.AC.UK 
Sent: Thursday, 14 March 2013, 10:27
Subject: [ccp4bb] PDBe website update: What's new
 

Dear all,

Twice a year, the Protein Data Bank in Europe (PDBe; http://pdbe.org) releases 
new and improved tools and services. Our first website update of 2013 features:

1. A new weekly overview of new biology in the PDB
2. Improved searching of EMDB entries with rapid filtering of
results
3. New features for visual analysis of EMDB entries
4. Improved pages with chemical-shift-validation information for NMR
entries
5. Easy sharing of PDBe pages



1. A new weekly overview of new biology in the PDB.

You can now view instances of new biology in the PDB including Pfam
families, GO terms and UniProt entries that appear in the archive
for the first time. For example, this week’s release includes the
first structure ever from a member of the Alternative Oxidase Pfam
sequence family
(http://www.ebi.ac.uk/pdbe/searchResults.html?display=latesttab=xref). 
Note that two kinds of new biology are shown: cases where an existing Pfam 
family etc. maps to a newly released structure in the PDB for the first time, 
and cases where an existing PDB entry maps to a new entry in Pfam, GO, UniProt, 
CATH or SCOP.

2. Improved searching of EMDB entries with rapid filtering of
results.

We have made searching the EMDB easier by implementing search
filters that enable you to rapidly narrow down the set of search
results. Filters based on experimental details, journal, organism
etc. can be applied when searching (http://pdbe.org/emsearch) or browsing 
the contents of the archive (http://pdbe.org/embrowse).

3. New features for visual analysis of EMDB entries.

We have introduced several new features to aid visual analysis and
validation of EMBD entries, including:

* A volume-estimate graph that shows how the enclosed volume of a map 
varies with the contour level (e.g. http://pdbe.org/emd-5357/analysis).
* A Fourier-shell correlation (FSC) curve used to estimate the 
resolution of single particle maps (e.g. http://pdbe.org/emd-5357/analysis).
* A residue-based atom-inclusion chart that can be used to assess the 
quality of fit of PDB models to an EMDB map (e.g.
* http://pdbe.org/emd-2017/analysis).
* An image showing the overlay of any deposited masks on a map, that 
show segmentations or some particular feature of an entry (e.g. 
http://pdbe.org/emd-1206/analysis).
(Note that not all features are available or applicable for all entries.)

4. Improved pages with chemical-shift-validation information for NMR
entries.

We have improved the presentation of information on the validation
of chemical shifts with VASCO. The redesigned webpages now show
statistics on the assigned chemical shifts, any referencing
corrections and a list of atoms with unusual chemical shifts values
(e.g. http://www.ebi.ac.uk/pdbe-apps/nmr/vasco/searchEntry?pdbCode=2knr).

5. Easy sharing of PDBe pages.

Regular visitors of the PDBe website may have noticed some small
changes to the headers of our pages. The new headers include
feedback and sharing buttons on every PDBe webpage. If you want to
share what you find on one of our pages with friends or colleagues,
use the share button to post the URL on Facebook, Twitter, etc. or
to email it. If you have suggestions for improvements, please let us
know by using the feedback button.

You may also have noticed our new logo. Based on a motif that is
found at all levels of structure, we hope that our new logo will
quickly become synonymous with the provision of high‐quality
information about 3D molecular and cellular structure. Our new logo
and guidelines for its use are available from our website at 
http://pdbe.org/logo

As always, we welcome your comments and suggestions through the
feedback button at the top of every PDBe webpage.

Gary.


-- 
Gary Battle
Protein Data Bank in Europe (PDBe) http://www.facebook.com/proteindatabank 
http://twitter.com/PDBeurope 

Re: [ccp4bb] strict structure based alignment

[MARTYN SYMMONS]

Hi -  SSM algorithm at PDBeFold will do this sort of thing


http://www.ebi.ac.uk/msd-srv/ssm/

I wrote a tutorial for multiple alignment to go with a Phaser story

http://www.ebi.ac.uk/pdbe-apps/quips?story=Phaserauxpage=MultiplePDBeFoldminitutorial

The last page of this tells you where to get an multiple fastA alignment of the 
superimposed structures and explains the format. 


Hope that helps.
  Martyn 


Martyn Symmons

EBI Cambridge

    





 From: Eric Pettersen p...@cgl.ucsf.edu
To: CCP4BB@JISCMAIL.AC.UK 
Sent: Monday, 16 July 2012, 2:17
Subject: Re: [ccp4bb] strict structure based alignment
 

On Jul 13, 2012, at 4:00 PM, Christian Roth wrote:

I want align a couple or protein structures by secondary structure matching to 
one target and want get a kind of aminoacid alignment file e.g. what residue 
fit 
the other, without adjustments due to sequence based alignments. 
I tried Strap, but as far as I understood it, it takes also the sequence into 
account. I tried also Rapido, but this does only a pairwise comparison. 
Superpose does align it nicely (ccp4 based or Coot based) but there seems to 
be no option to print the sequence alignment in a file and it is again  just a 
pairwise comparison .
Is there an other program which does something similar?

If you use UCSF Chimera (www.cgl.ucsf.edu/chimera), you can use the MatchMaker 
tool to superimpose the structures.  MatchMaker allows you to adjust the weight 
of sequence similarity vs. secondary structure matching, so you can just make 
the sequence similarity 0% and the secondary structure 100%.  With the 
structures superimposed, you can use the Match-Align tool to generate a 
sequence alignment based solely on the proximity of residues to one another in 
space.  Be warned that Match-Align will be very slow for 10+ structures, but 
is fine for half a dozen or so.  The generated alignment will be displayed in a 
window that will have a File menu where you can save the alignment to a 
variety of common formats.

--Eric

Eric Pettersen
UCSF Computer Graphics Lab

[ccp4bb] studentship posting

[MARTYN SYMMONS]

Posted on behalf of Dr Callaghan - please contact her directly.


4 year Ph.D. Studentship on RNase structure and metabolic control.
School of Biological Sciences.
University of Portsmouth.

Supervisor: Dr Anastasia Callaghan 

Co-supervisor: Dr Sarah Newbury (Sussex Medical School)

Understanding how metabolism is controlled within a cell is fundamentally 
important and is directly applicable to medical, environmental and 
biotechnological advances. 

Our studies have recently identified that a molecule of central metabolism 
interacts with an RNase and affects its ability to destroy mRNA. This project 
will unravel the details of this newly discovered mechanism and investigate 
whether it represents a conserved metabolite-RNase communicative link in 
prokaryotes and eukaryotes. 

Training will be provided in a range of molecular biology, biochemical, 
biophysical, and structural techniques. 

Please see: www.sebnet.org.uk  www.findaphd.com

Informal enquires welcome: anastasia.callag...@port.ac.uk

Application deadline: 31st January 2012

Re: [ccp4bb] [off topic] Control of crystals' direction and position in the drop.

[MARTYN SYMMONS]

Hi there 

 one thing I did to get more space for needles in a drop was to make 
'book end' coverslips. Your take a plastic covership and with a razor blade you 
cut obliquely into the plastic to make a 'flap' of the plastic. Then you lever 
it up to vertical to give a projecting vertical surface. You can then apply the 
drop to the angle between the flap and the coverslip. This supports the drop so 
that it does not flatten so much during equilibration. Making the cut takes a 
bit of trial and error but the plastic coverslips are not that expensive. It 
certainly gave me some chunkier needles and gave them space to grow upwards. 
Otherwise they flattened down as the droplet flattened with equilibration and 
often they 'glued' themselves to the horizontal surface.


    Hope it helps
    Martyn 


Martyn Symmons
Cambridge




From: Frederic VELLIEUX frederic.velli...@orange.fr
To: CCP4BB@JISCMAIL.AC.UK
Sent: Monday, 14 November 2011, 22:24
Subject: Re: [ccp4bb] [off topic] Control of crystals' direction and position 
in the drop.

Hi,

Your email mentions drop.

What about trying another technique where you do not have drops, such as the 
liquid interface diffusion method (in capillaries), or the use of dialysis ? 
Crystallisation 
under oil (injection of the 2 components under oil) could also be tried.

Fred

 Message du 14/11/11 22:15
 De : Nian Huang 
 A : CCP4BB@JISCMAIL.AC.UK
 Copie à : 
 Objet : [ccp4bb] [off topic] Control of crystals' direction and position in 
 the drop.
 
 Dear All,
 Does anybody find a way to control a crystal's positioning in the drop? I
 have needle shaped crystals. What I found out is that the vertical
 positioned crystals always grow much thicker than the crystals laying
 flatly. But unfortunately, it is a completely random event and only 1%
 crystals can appear vertically. I have tried different formats of plates
 and equilibration techniques w/o much success. Any suggestion is highly
 appreciated.
 
 Nian Huang, Ph.D.
 UT Southwestern Medical Center
 

Re: [ccp4bb] [off topic] Control of crystals' direction and position in the drop.

[MARTYN SYMMONS]

Attached is a picture of making the bookend setup - step b. shows the razor 
levering up the flap from the plastic coverslip  - be careful not to cut right 
through (I have done that and the drop dries extra quick in such a case) also 
be careful to apply the protein/well droplet as shown with to the shiny side of 
the lifted flap. (Apologies for the attachment). 

hope your crystals go well.

Martyn 


From: Nian Huang huangn...@gmail.com
To: CCP4BB@JISCMAIL.AC.UK
Sent: Tuesday, 15 November 2011, 17:44
Subject: Re: [ccp4bb] [off topic] Control of crystals' direction and position 
in the drop.


Thank you guys. I basically tried almost everything that I can find in the 
hampton catalogue and in this bulletin, seeding, hanging, sitting, sandwitch 
drops (which made things worse), temperature, gel, oil batch, and additive 
screens. Manipulating the crystal with fiber increases the chance to make it 
grow twin. The book end coverslip sounds a fantasic idea. It might be just 
going to work.

Best,

Nianattachment: bookend.jpg

Re: [ccp4bb] Protein sequencing service?

[MARTYN SYMMONS]

I have had excellent service from Mike Weldon (m.a.wel...@bioc.cam.ac.uk) at 
PNAC University of Cambridge Biochemistry. He will accept outside orders 
subject to capacity being available.

http://www.bioc.cam.ac.uk/pnac/

Cheers
  Martyn 

Martyn Symmons
EBI - Cambridge
 
 On 24/09/2010 4:46, Rex Palmer wrote:
  Can anyone please recommend a UK protein sequencing
 service. Our protein
  is dimeric with reported molecuar weight of about
 85kDa.Our funds are
  somewhat limited.
  Thanks in advance.
  Rex Palmer
  Birkbeck College, London
  RexPalmer2010.homestead.com
 
 --         Andreas Förster,
 Research Associate
         Paul Freemont  Xiaodong
 Zhang Labs
 Department of Biochemistry, Imperial College London
             http://www.msf.bio.ic.ac.uk

Re: [ccp4bb] protein turns brown

[MARTYN SYMMONS]

and possibly the opposite to reduction which is oxidation - do you have cys 
residues? Perhaps your DTT or TCEP got exhausted? Remember you can add up to 
10mM (not the traditional token 1mM). But remember to neutralize TCEP. Other 
possibility is adventitious metals as these give strong charge transfer bands 
on binding to cys.

So make sure you include EDTA or similar chelator (after metal affinity 
obviously). Oxidation is more likely if your protein unfolds - so try to keep 
it happy with nicely buffered pH and any ligands that might stabilize it.
   Good luck. 
 Martyn
Martyn Symmons
Cambridge

--- On Fri, 24/9/10, vikrant saa powervikr...@yahoo.co.in wrote:

From: vikrant saa powervikr...@yahoo.co.in
Subject: Re: [ccp4bb] protein turns brown
To: CCP4BB@JISCMAIL.AC.UK
Date: Friday, 24 September, 2010, 10:50


sometime it does happen becoz of protein aggregation, or reducing environment.
but it may be your protein color as well that visible during concentration 
 
 
Vikrant


 
 
 






From: sandeep toskgu...@rediffmail.com
To: CCP4BB@JISCMAIL.AC.UK
Sent: Fri, 24 September, 2010 2:04:54 PM
Subject: [ccp4bb] protein turns brown

Dear all,

I have purified protein from E.coli. expression system. the protein has been 
purified with three independant columns. Now during concentration step using 
amicon, the protein shows brown colour. what could be the reason.

best regards and Thanks,
sandy 

Re: [ccp4bb] Effect of NCS on estimate of data:parameter ratio

[MARTYN SYMMONS]

Dear All
one thing I remembered from what Gerard pointed out was the difference 
in the XPLOR/CNS formalism between strict and restrained which is not a 
continuum. Restrained was obviously when you had multiple copies and they were 
restrained with a weight (which was like a force constant) to be similar when 
superimposed. So if you increase the force constant then they can move during 
refinement but they all try to move together when they move. 

And the other extreme is strict where there was no force applied at all but 
only a single copy of the chain and the ASU is built by applying the NCS 
symmetry. The atoms are free to move but, unlike the case with restrained where 
there is superimposition on the fly, in the strict case there is no automatic 
update of the superimposition matrices. So every move gets religiously copied 
to all the chains when the ASU is made. At this point I guess the copies can 
bump and so apply a force on each other but that is a local, and likely to be 
perturbing, force. 

best wishes
 Martyn 

Martyn Symmons
Cambridge



--- On Thu, 23/9/10, Ian Tickle ianj...@gmail.com wrote:

 From: Ian Tickle ianj...@gmail.com
 Subject: Re: [ccp4bb] Effect of NCS on estimate of data:parameter ratio
 To: CCP4BB@JISCMAIL.AC.UK
 Date: Thursday, 23 September, 2010, 11:21
 Hi Gerard  Pavel
 
 Isn't this the proviso I was referring to, that one cannot
 in practice
 use an infinite weight because of rounding errors in the
 target
 function.  The weight just has to be 'big enough' such
 that the
 restraint residual becomes sufficiently small that it's no
 longer
 significant.
 
 In numerical constrained optimisation the method of
 increasing the
 constraint weights (a.k.a. 'penalty coefficients') until
 the
 constraint violations are sufficiently small is called the
 'penalty
 method', see http://en.wikipedia.org/wiki/Penalty_method .  The
 method
 where you substitute some of the parameters using the
 constraint
 equations is called (you guessed it!) the 'substitution
 method', see
 http://people.ucsc.edu/~rgil/Optimization.pdf . 
 There are several
 other methods, e.g. the 'augmented Lagrangian method' is
 very popular,
 see 
 http://www.ualberta.ca/CNS/RESEARCH/NAG/FastfloDoc/Tutorial/html/node112.html
 .  As in the penalty method, the AL method adds
 additional parameters
 to be determined (the Lagrange multipliers, one per
 constraint)
 instead of eliminating some parameters using the constraint
 equations;
 however the advantage is that it removes the requirement
 that the
 penalty coefficient be very big.
 
 The point about all these methods of constrained
 optimisation is that
 they are in principle only different ways of achieving the
 same
 result, at least that's what the textbooks say!
 
 And now after the penalties and substitutions it's time to
 blow the whistle ...
 
 Cheers
 
 -- Ian
 
 On Wed, Sep 22, 2010 at 10:00 PM, Pavel Afonine pafon...@lbl.gov
 wrote:
   I agree with Gerard. Example: it's unlikely to
 achieve a result of
  rigid-body refinement (when you refine six
 rotation/translation parameters)
  by replacing it with refining individual coordinates
 using infinitely large
  weights for restraints.
  Pavel.
 
 
  On 9/22/10 1:46 PM, Gerard DVD Kleywegt wrote:
 
  Hi Ian,
 
  First, constraints are just a special case of
 restraints in the limit
  of infinite weights, in fact one way of
 getting constraints is simply
  to use restraints with very large weights
 (though not too large that
  you get rounding problems). These
 'pseudo-constraints' will be
  indistinguishable in effect from the 'real
 thing'.  So why treat
  restraints and constraints differently as far
 as the statistics are
  concerned: the difference is purely one of
 implementation.
 
  In practice this is not true, of course. If you
 impose infinitely strong
  NCS restraints, any change to a thusly restrained
 parameter by the
  refinement program will make the target function
 infinite, so effectively
  your model will never change. This is very
 different from the behaviour
  under NCS constraints and the resulting models in
 these two cases will in
  fact be very easily distinguishable.
 
  --Gerard
 
 
 **
                            Gerard J.
  Kleywegt
    Dept. of Cell  Molecular Biology
  University of Uppsala
                    Biomedical Centre  Box
 596
                    SE-751 24 Uppsala
  SWEDEN
 
     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.
 
 **
 

Re: [ccp4bb] PEG in the pdb?

[MARTYN SYMMONS]

Dear Klaus
 depends how many missing residue you want advertised in the 
header of your deposited file. The pdb will put in a special Remark 610 and 
Remark 615 to indicate any missing residues from your heterogen list. However 
the policy is not to list every missing (or zero occupancy) atom. 
  I have fitted fragments of pegs into density - so I know there are defined 
hetero residues for several sizes of small fragments. But if you name a PEG as 
a larger fragment than you can see then, as each PEG would be only one residue, 
effectively you get only one line in a remark 610 for each PEG with missing 
bits. 
  Zero occupancy is generally a deprecated way of dealing with missing density 
as it is confusing for less experienced user of the coordinates. I think zero 
occupancy can be useful during refinement as the atoms help fill space (or for  
example satisfy NCS restraint format requirement) but then these atoms can be 
stripped out before deposition. They should in any case never be included in 
B-factor refinement as they will skew the statistics and possibly the B-factor 
restraint model.   
  It seems that large numbers of missing heterogen atoms were anticipated when 
the format was drawn up - hence the absence of a requirement to list all 
heterogen atoms that are missing. 
  Hope that helps. 
    Martyn 

EBI, Cambridge



--- On Thu, 12/8/10, Klaus Sengstack sengstack-kl...@yahoo.de wrote:

From: Klaus Sengstack sengstack-kl...@yahoo.de
Subject: [ccp4bb] PEG in the pdb?
To: CCP4BB@JISCMAIL.AC.UK
Date: Thursday, 12 August, 2010, 9:16

Hi everybody,

I just solved the structures of an enzyme an some variants. In the active site 
cavity of each variant I found one or two fragments of PEG1000 bound. I used 
PEG1000 in the crystallization condition. Among the enzyme variants the number 
of non-hydrogen atoms of these PEG fragments varies between 7 and 19 atoms. Now 
I want to deposit the structures in the pdb and my question is, if I have to 
define each fragment as a single ligand (what would be a lot of work) or can I 
define them as PEG1000 molecules? Thanks.

K.S.

Re: [ccp4bb] PEG in the pdb?

[MARTYN SYMMONS]

That's a good point, Ed

Based on the formula: HO CH2-(CH2-O-CH2)n-CH2OH, PEG MW = 44n+62, a table of n 
to MW goes like below which gives an idea of what range is possible. Someone 
maybe knows what polydispersity can be expected from the synthetic process. As 
you say though, a specific range could partition out of the bulk to the protein 
surface. And the main point of the original post was that only a subset of 
atoms in the bound PEG may be ordered enough to see. 
 n   MW
 5, 282.
 6, 326.
 7, 370.
 8, 414. 
 9, 458. 
 10, 502.
 11, 546.
 12, 590.
 13, 634.
 14, 678.
 15, 722.
 16, 766.
 17, 810.
 18, 854.
 19, 898. 
 20, 942.
 21, 986. 
 22, 1030. 
 23, 1074. 
 24, 1118.
 25, 1162.
 26, 1206. 
 27, 1250. 
 28, 1294. 
 29, 1338. 
 30, 1382.
 31, 1426. 
 32, 1470. 
 33, 1514. 
 34, 1558.
 35, 1602.
 36, 1646.
 37, 1690.
 38, 1734.
 39, 1778.
 40, 1822.
 41, 1866. 
 42, 1910. 
 43, 1954.
 44, 1998.
 45, 2042.
 46, 2086.
 47, 2130. 



--- On Thu, 12/8/10, Ed Pozharski epozh...@umaryland.edu wrote:

 From: Ed Pozharski epozh...@umaryland.edu
 Subject: Re: [ccp4bb] PEG in the pdb?
 To: CCP4BB@JISCMAIL.AC.UK
 Date: Thursday, 12 August, 2010, 17:35
 PEG solutions contain fragments of
 all sizes - it is the average size
 (however defined by the manufacturer) that is 1000. 
 So technically it
 is incorrect to claim that you have PEG1000 molecules bound
 to your
 protein, it is most likely much shorter fragments that can
 penetrate the
 channels in protein crystals.
 
 It's not a lot of work to generate monomer libraries for
 peg fragments
 of different length (and some are available from standard
 monomer
 libs).  
 
 I always wondered why PEG is not defined in the standard
 libraries as a
 polymer - perhaps because it is rarely needed.  Or is
 it?
 
 Ed.
 
 On Thu, 2010-08-12 at 08:16 +, Klaus Sengstack wrote:
  Hi everybody,
  
  I just solved the structures of an enzyme an some
 variants. In the
  active site cavity of each variant I found one or two
 fragments of
  PEG1000 bound. I used PEG1000 in the crystallization
 condition. Among
  the enzyme variants the number of non-hydrogen atoms
 of these PEG
  fragments varies between 7 and 19 atoms. Now I want to
 deposit the
  structures in the pdb and my question is, if I have to
 define each
  fragment as a single ligand (what would be a lot of
 work) or can I
  define them as PEG1000 molecules? Thanks.
  
  K.S.
  
  
 
 -- 
 I'd jump in myself, if I weren't so good at whistling.
                
            
    Julian, King of Lemurs

Re: [ccp4bb] How to distinguish microcrystalline, quasicystalline and precipitation?

[MARTYN SYMMONS]

Dear Joy
  - microcrystalline in most cases would be birefringent - this is 
difficult to observe these days in our plastic boxed crystallization world with 
colours everywhere. 

But if you really want to know you can pipette the droplet onto a glass 
coverslip and dim the room lights. Set the polarizers _exactly_ to extinction. 
Then you can see if the precipitate glows. 

Of course it could be a cubic space group crystal which I heard does not show 
BR.

And it could be micro-spherulites which are indeed quasicrystalline - these are 
strongly bireringent - maybe _too_ strongly is the clue here! If any get to a 
reasonable size then you can see a X-like pattern in the centre of these since 
they appear to be ordered in spherical shells of unfolded protein. 

One other clue can be the density - most protein crystals are not far off the 
droplet density. So if the precipitate settles strongly into a circle at the 
bottom of the droplet then I would say this is a bad sign :(

Hope this helps.

regards
    Martyn Symmons

EBI, Hinxton, Cambs. UK 

 

--- On Thu, 10/6/10, joybeiyang joybeiy...@gmail.com wrote:

From: joybeiyang joybeiy...@gmail.com
Subject: [ccp4bb] How to distinguish microcrystalline, quasicystalline and 
precipitation?
To: CCP4BB@JISCMAIL.AC.UK
Date: Thursday, 10 June, 2010, 7:06



 
 
#yiv1007907758 BLOCKQUOTE {
MARGIN-TOP:0px;MARGIN-BOTTOM:0px;MARGIN-LEFT:2em;}
#yiv1007907758 OL {
MARGIN-TOP:0px;MARGIN-BOTTOM:0px;}
#yiv1007907758 UL {
MARGIN-TOP:0px;MARGIN-BOTTOM:0px;}

 
Hi everyone, I am preparing a crystallization 
manual for our group, however, I found that it is very difficult to 
distinguish 
microcrystalline, quasicrystalline and precipitation, especially when the 
precipitation was shiny, like the grit on the beach. Is there a way to 
distinguish the three? 
 
Comments and suggestions will be greatly 
appreciated!




Joy

[ccp4bb] Fw: [ccp4bb] degradation of protein durring freez thaw

[MARTYN SYMMONS]

Sounds like good advice - I suppose that since pure water un-freezes last, you 
could have protein concentration inhomogeneities? So in areas where the protein 
concentration is raised it could start to feel like crashing out.


All the best
Martyn

Martyn Symmons
Cambridge



- Original Message 
From: Frank Niesen dr.frank.nie...@gmail.com
To: CCP4BB@JISCMAIL.AC.UK
Sent: Friday, 23 April, 2010 10:40:59
Subject: Re: [ccp4bb] degradation of protein durring freez thaw

An aspect that is sometimes overlooked is that the need to avoid mini 
freeze-thaw cycles does not only call for a quick, snap-freeze process for 
samples (in thin-walled PCR tube and not too large volume; 50-100 ul I'd 
recommend), but - in turn - also for quick thawing: I prefer holding the PCR 
tube under running cold water straight after removal from the freezer, and 
stick into the ice bucket only after I see that all of the sample is thawed.
Best,
Frank

Re: [ccp4bb] Phasing statistics

[MARTYN SYMMONS]

Sorry - I had not twigged that this was a SAD discussion. 

In this case DM is as you say gonna save you from bimodal ambiguities - as we 
all know that is why the FOMs get so much better during DM - but that seems 
fine to me as it is pretty much new information coming in from the solvent 
flattening, histogram matching, probably some averaging. 

So that improvement is not 'artifactual' but in fact part of the experimental 
detail of just how the structure was solved. I guess FOMs are really only 
useful in the Crick and Blow centroid case.

Still think any and all info on the experiment(s) is useful to see - if only 
for people to plan their own experiments building on past experience.

Best wishes and regards
Martyn 

Martyn Symmons
Cambridge

 



- Original Message 
From: Ian Tickle ianj...@gmail.com
To: CCP4BB@JISCMAIL.AC.UK
Sent: Wednesday, 14 April, 2010 15:42:51
Subject: Re: [ccp4bb] Phasing statistics

I have to say that I don't share James' enthusiasm for the FOM as a
useful statistic, even for experimental phases, and particularly not
in the SAD (and SIR) cases, which after all is what this thread is
supposed to be about.   This is even without delving further into the
murky issues of wildly inflated FOM estimates from DM that he and
others have raised.

My point is that the definition of FOM is that it's the *expected*
cos(phase error), as opposed to *actual* cos(phase error) and the
'expected' bit makes all the difference!  It's essentially the
difference between precision and accuracy, i.e. the FOM tells you how
precise the phase estimate is and not at all how accurate it is: for
the latter you need to calculate the errors relative to the phases of
the final model, as James suggested.  The FOM is roughly related
(anti-correlated to be precise) to the variance: this is clear if we
take the case of small phase deviations, then cos(x) expands to
(1-x^2/2) and the variance is x^2, where x is the deviation from
the mean (NOT the same thing as the error!).  So just like the
variance it measures the 'peakedness' of the distribution: a FOM=1
corresponds to variance=0, i.e. an infinitely sharp distribution
(delta function).

Now a particular problem arises in SAD (and SIR) because then in
general we always get a bimodal distribution: i.e. 2 separate peaks.
If these peaks happen to separated by 180 deg then the FOM is 0
(cos(90)=0).  A large separation is most likely to occur when the
anomalous contribution is large, when of course you expect the phase
estimates to be optimal, for example see Fig 2(a) in Wang et al., Acta
Cryst. (2007). D63, 751–758.  Conversely if the anomalous contribution
is small, so the phase estimates are poor, the separation between the
2 estimates is small and the FOM is close to 1!  So we apparently have
a situation where the *better* are the phases, the *lower* is the FOM!
What the Figure in the Wang paper ignores of course are the
experimental errors which will tend to broaden the distribution and
lower the FOM in the case where the anomalous contribution is small
relative to the errors.  Even so it means that the FOM is not a good
measure of phase quality.  Much better IMO is the phasing power (mean
heavy-atom amplitude / mean P-weighted lack of closure error).  This
essentially measures the degree to which the phase ambiguity is
capable of being successfully resolved by DM methods.

One other thing puzzles me: why do many programs that purport to
calculate average phase differences (relative to model phases say) use
statistics such as, well, average |phase difference| when we already
have a perfectly good measure in the FOM!, i.e. average cos(phase
difference)?  Then you would be able to directly compare the FOMs from
experimental phases with those from model phases.  Even better still
would be the average log likelihood: if the phase probability is exp(A
cos(delta_phi)) then the log likelihood is simply A cos(delta_phi) and
you just average that, i.e. it's essentially the averaged FOM-weighted
cos(phase difference), so that the average is weighted according to
the reliability of the phase estimates.  A poor phase estimate is
likely to be associated with a small F which is not going to
contribute much to the map anyway, so it makes no sense to give poor
phases the same weight as good phases in the average.

Cheers

-- Ian

On Tue, Apr 13, 2010 at 6:47 PM, James Holton jmhol...@lbl.gov wrote:
 Probably the only phasing stat that I pay any attention to these days is the
 Figure of Merit (FOM). This is because, the _definition_ of FOM is that it
 is the cosine of the phase error (or at least your best estimate of it).
  FOM=1 is perfect phases and FOM=0 is random phases, and a reasonable cutoff
 value for FOM is 0.5 (see Lunin  Woolfson, Acta D, 1993).  Yes, there are
 ways to get various programs to report very inaccurate values for FOM (such
 as running DM for thousands of cycles), and yes, there are often legitimate
 reasons to run these programs in this way

Re: [ccp4bb] Phasing statistics

[MARTYN SYMMONS]

I agree this is very interesting -

The 'real' FOM is a great idea (we should start a campaign for real FOMs!) 

What the 'real' FOM is trying to get at is the quality of the initial 
experimental map. This could also be subsequently calculated for NCS-averaged 
and density-modified maps subsequently to show how the building was achieved.

 Gamma correction and real space free residuals can be used to avoid 
overfitting (DM has these I know, prob other progs too). But the point that 
overfitting can help the crystallographer build a correct structure is a good 
one. Conversely I have heard of structures that are not just correct but also 
wonderfully original, built by talented crystallographers into initially 
apparently difficult-to-interpret maps. So we maybe need to celebrate what can 
be done; not use this as necessarily a criticism of the final structure.

One of the things I heard from F. VonD was 'don't get hung up on statistics - 
look at maps'. Any map is going to have ropey bits as well as easy bits - so 
the only way to convey that would not be one number like a FOM.  But more like 
a residue-by-residue real-space Rfactor of the _final_ model in the _original_ 
map?

In the real world ;) I don't see that happening anytime soon

all the best
   Martyn

Martyn Symmons
Cambridge





- Original Message 
From: Soisson, Stephen M stephen_sois...@merck.com
To: CCP4BB@JISCMAIL.AC.UK
Sent: Tuesday, 13 April, 2010 19:09:12
Subject: Re: [ccp4bb] Phasing statistics

This is an interesting thread, and perhaps I should not dive in on such
a heady topic, BUT, I do want to point out my own particular bias
regarding FOM that is not entirely consistent with James' point of view.
In my experience, the FOM obtained after density modification runs are
almost always extremely optimistic - I have seen relatively high
apparent FOM after density modification runs (0.7) that had nearly
uninterpretable maps. As such, I am much more interested in knowing
about the FOM from the phasing calculations themselves and NOT after
density modification.  

That said, applying arbitrary cut-offs to what would be deemed an
acceptable FOM, after phasing calculations, to generate maps that are
interpretable is not really a good thing to do.  For instance, I just
had a structure where the FOM was 0.35 after phasing (a rubbish
structure perhaps?), BUT, the data are highly redundant and the solvent
content in the high 70% range.  The post density modified maps are
stunningly good.  One could easily imagine many other scenarios (e.g.
NCS) where the modified maps and apparent FOM would be decoupled.

So, I do agree with James's suggestion that perhaps we should be
retrospectively calculating a real FOM between the final model and the
actual maps you built into (after whatever you did to get them).  This
seems like a very good idea indeed.

More personal biases revealed:  I actually look at, and use, the Cullis
R values on anomalous and isomorphous data to help determine how much
signal is in the data.  Simplified, these numbers are your Average
estimated lack-of-closure divided by your average observed difference.
That's an important thing to know, and I find them quite useful.

Steve

-Original Message-
From: CCP4 bulletin board [mailto:ccp...@jiscmail.ac.uk] On Behalf Of
James Holton
Sent: Tuesday, April 13, 2010 1:48 PM
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] Phasing statistics

Probably the only phasing stat that I pay any attention to these days is

the Figure of Merit (FOM). This is because, the _definition_ of FOM is 
that it is the cosine of the phase error (or at least your best estimate

of it).  FOM=1 is perfect phases and FOM=0 is random phases, and a 
reasonable cutoff value for FOM is 0.5 (see Lunin  Woolfson, Acta D, 
1993).  Yes, there are ways to get various programs to report very 
inaccurate values for FOM (such as running DM for thousands of cycles), 
and yes, there are often legitimate reasons to run these programs in 
this way.   But, there are also very wrong things one can do to get low 
Rmerge, Rcryst, and especially Rfree.  It is simply a matter of knowing 
(and reporting) what you are doing.


If you are worried that your favorite estimate of FOM is inaccurate, 
then you can always turn to your most accurate phases:  those of your 
final, refined model (the one that you have convinced yourself is 
right and ready to publish).  Taking these as the true phases, the 
true FOM can always be obtained by comparing the final-model phases to

those of your initial map (using PHISTATS or SFTOOLS).  This is by no 
means standard practice, but perhaps it should be?


Anyway, FOM is _supposed_ to be the cosine of the phase error, and is 
therefore the most relevant statistic when it comes to how good your 
phases were when you started building.  This is why it is important as a

reviewer to know what it is.  If I am faced with a structure that was 
built into a MAD map with initial FOM = 0.8 to 2

Re: [ccp4bb] Phasing statistics

[MARTYN SYMMONS]

Hiya Frank

Well my take on it would be that they did a certain experiment - which includes 
phasing datasets - and we have an expectation to see all the steps reported. 
Good structures are not the 'be all' and 'end all'. Reporting data is not 
merely to convince readers that this is not a 'made up' structure. It is like 
in mathematics when you don't just report the correct answer but your 'show 
your workings' as my old maths teacher used to say. So maybe phasing statistics 
are pretty important in that respect. 

And it would be good to have datasets with experimental phases deposited. I 
should've done this with my 1e3h years ago as the experimental phases (which, 
as I recall, you were a friendly advisor on ;) were included in the refinement 
target (CNS mlhl). SeMet phases are usually good - but ones from other heavy 
atoms it might be nice to see too. There used to be a heavy atom database for 
such like things - not sure of the current status of that.

see you
 all the best
   Martyn 

Martyn Symmons
Cambridge












From: Frank von Delft frank.vonde...@sgc.ox.ac.uk
To: CCP4BB@JISCMAIL.AC.UK
Sent: Monday, 12 April, 2010 12:24:50
Subject: Re: [ccp4bb] Phasing statistics

I fully agree, for high quality data.

What though if the data are not impeccable and the structure
necessarily ropey?  E.g. 4A phases and anisotropic diffraction.  By
what metrics do we then judge the results?

(I don't know the answer, btw, but our membranous colleagues surely
spend quite a bit of time with that question...)

phx.


On 12/04/2010 12:10, Anastassis Perrakis wrote: 
Hi -
 


A year or so ago, I have asked as a referee somebody to provide
for a paper the statistics for their heavy atom derivative dataset,
 
and for the phasing statistics. For some good reasons, they were
unable to do that, and they (politely) asked me
'what would it change if you knew these, isn't the structure we
present impeccable?'. Well, I think they were right.
Their structure was surely correct, surely high quality. After
that incident and giving it some thought, 
I fail to see why should one report e.g. PP or Rcullis, or why
will I care what they were if the structure has a convincing Rfree and
is properly validated. 
If someone wants to cheat at the end of the day, its easy to
provide two numbers, but its hard to provide a good validated model
that agrees with the data.
(and, yes, you can also make up the data, but we have been
there, haven't we?!?)


So, my question to that referee, likely being a ccp4bb
aficionado that is reading this email, or to anyone else really, is:


What would it help to judge the quality of the structure or the
paper if you know PP, Rcullis and FOM?


Best -


A.


PS Especially since you used SHELXE for phasing these statistics
are utterly irrelevant, and possibly you could advice the referee to
read a bit about how SHELXE works ... or go to one of the nice courses
that George teaches ...


On Apr 12, 2010, at 10:37, Eleanor Dodson wrote:

You can feed the SHELX sites into phaser_er or CRANK both of
which will 
give this sort of information.

Or mlphare if you know how to set it up..

Eleanor


Harmer, Nicholas wrote:

Dear CCP4ers,



I've been asked by a referee to provide the
phasing statistics for a SAD dataset that I used to solve a recent
structure. Whilst I have been able to find a figure-of-merit for the
data after phasing, I can't work out how to get any other statistics
(e.g. phasing power or an equivalent or Rcullis). Does anyone know a
good route to obtaining useful statistics to put in the paper for SAD
data?



The structure solution was carried out
using SHELX C/D/E and then ARP/wARP.



Thanks in advance,



Nic Harmer



=

Dr. Nic Harmer

School of Biosciences

University of Exeter

tel: +44 1392 725179




P please don't print this e-mail unless you really need to
Anastassis (Tassos) Perrakis, Principal
Investigator / Staff Member
Department of Biochemistry (B8)
Netherlands Cancer Institute, 
Dept. B8, 1066 CX Amsterdam, The Netherlands
Tel: +31 20 512 1951 Fax: +31 20 512
1954 Mobile / SMS: +31 6 28 597791

Re: [ccp4bb] 2,3-butanediol

[MARTYN SYMMONS]

One thing I found with 2,3-Butanediol was for some crystallizations you can 
make it up a cryo solution with it at 12% and then put a thin layer it on top 
of the crystal droplet in a sitting drop well. Then you can loop the crystal 
and pull it up through the cryo layer - then out and to N2(l).  Just how brief 
this is depends on your dexterity and also how well the crystal is engaged in 
the loop. My crystals were not very happy about the cryo but I could make it 
brief enough to get them out intact.

Good luck,

Martyn 

Martyn Symmons
MBU Cambridge




- Original Message 
From: Sean Seaver s...@p212121.com
To: CCP4BB@JISCMAIL.AC.UK
Sent: Wednesday, 24 March, 2010 18:47:38
Subject: Re: [ccp4bb] 2,3-butanediol

Dear all,
I would like to know which is the more suitable method of using 2,3-Butanediol 
as cryoprotectant? Addition in to reservoir buffer or crystal soaking or both 
can be tried?

Cheers
Gauri
--
I am sure both could be tried, but have found quite few papers mentioning brief 
soaking with 2,3-butanediol:

Purification, crystallization and X-ray diffraction analysis of the 
extracellular part of the human Fc receptor for IgA, Fc[alpha]RI (CD89)
http://scripts.iucr.org/cgi-bin/paper?cnor=bw5002
5-10 seconds

Purification, crystallization and X-ray analysis of Hibiscus chlorotic ringspot 
virus
http://scripts.iucr.org/cgi-bin/paper?cnor=gr2202
described as 'briefly'

Crystallization and preliminary X-ray analysis of recombinant human acid 
[beta]-glucocerebrosidase, a treatment for Gaucher's disease
http://scripts.iucr.org/cgi-bin/paper?cnor=pu0086
described as 'briefly'

Crystallization and preliminary X-ray crystallographic studies of recombinant 
human betaine-homocysteine S-methyltransferase
http://scripts.iucr.org/cgi-bin/paper?cnor=gr2115
'quickly transferred'

Purification, crystallization and X-ray analysis of Hibiscus chlorotic ringspot 
virus
http://scripts.iucr.org/cgi-bin/paper?cnor=pu0115
5% increments from 5 to 30% for 3 min at each step

Hope that helps,

Sean

Re: [ccp4bb] crystallization of a macromolecular complex

[MARTYN SYMMONS]

I agree this 3:1, 4:1, ... 10:1 etc is a good approach - especially for 
screening - as it samples more concentration conditions than the plain vanilla 
1:1 set-up. One generally good thing too is that the precipitant also starts at 
lower concentration - so less chance of setting up a load of trials and 
watching them immediately crash out (although obviously not a problem in this 
particular case!) .

I think the reason for differences from pre-prepared concentrated protein are 
often to do with other components that come in with the protein - so anything 
in the protein solution like buffer, salt, reductant, detergent, metal ions, 
etc will be concentrated up alongside the protein. 

cheers
Martyn 

Martyn Symmons, MBU Cambridge.

As an aside I heard that the 1:1 derives from the days when pipettes could not 
be trusted with such small volumes. The 1:1 ratio is the best chance to get a 
droplet with reproducible relative concentrations regardless of any systematic 
error in the exact volumes dispensed.

 







From: Jacob Keller j-kell...@md.northwestern.edu
To: CCP4BB@JISCMAIL.AC.UK
Sent: Friday, 5 March, 2010 17:02:15
Subject: Re: [ccp4bb] crystallization of a macromolecular complex

 

 
20 g/L is the same as 20 mg/ml, isn't it?  That does 
  not seem particularly high to me.
 
Why not try 200 g/L?
 
As a variant to 
this, you could just try doing 10:1 prot:ppt drops, and allowing the protein 
concentration to increase in situ. In my limited experience, however, this is 
not the same as setting up drops 1:1 with more concentrated protein, for 
reasons 
unknown to me. I wonder whether that is the experience of others on the BB as 
well?
 
JPK

Re: [ccp4bb] Per-residue RMSD for multiple structures

[MARTYN SYMMONS]

I did this recently for an analysis of the subunit variation in AcrB. I took 
the LSQMAN multi-RMSDs among the three chains after improved superimposition 
and edited them into the format that Rob Campbell's data2bfactor.py expected. 
And then I put them into the B-factor column of one subunit. Finally I invoked 
Rob Campbell's color_b.py in pymol to colour the molecular surface by them 
(Fig. 4 in http://www.pnas.org/content/106/17/7173/suppl/DCSupplemental). But I 
guess a backbone could be similarly. 

There are probably other ways to do this but Rob Campbell's scripts are a boon 
and a blessing

cheers,
Martyn 

Martyn Symmons
MRC-MBU



- Original Message 
From: Gerard DVD Kleywegt ger...@xray.bmc.uu.se
To: CCP4BB@JISCMAIL.AC.UK
Sent: Tuesday, 23 February, 2010 14:57:02
Subject: Re: [ccp4bb] Per-residue RMSD for multiple structures?

Thanks Stephen!

I was going to suggest that, but I was afraid of the self-appointed CCP4BB 
Gestapo that has been seen goose-stepping in this neighbourhood recently 
(Tassos recently accused me of becoming mellow and diplomatic in my dotage, so 
I hope I've set the record straight now). However, since this solution is 
neither CCP4 nor Phenix, we may get away with this heinous act of 
bulletin-board heresy... On the other hand, I've learned that it is often more 
expedient to beg for forgiveness than to ask for permission.

I would add that:

- I assume that the sequences and numbering are identical

- you should put the structures in one big PDB file and read it into LSQMAN

- since LSQMAN doesn't do true multiple-structure alignment, you could 
pre-align them, e.g. with SSM/PDBeFold

- if you didn't, you could indeed use the MCentral and MAlign commands to align 
them

- my favourite plot would be the CD plot (but then again, it would, wouldn't 
it?) - see for instance: 
http://xray.bmc.uu.se/cgi-bin/gerard/image_page.pl?image=usf/pics/cdplot_1ldn.gif
 - which is also produced with the MPlot command - 
http://xray.bmc.uu.se/usf/lsqman_man.html#S82 - a normal MPlot would look like 
this: 
http://xray.bmc.uu.se/cgi-bin/gerard/image_page.pl?image=usf/pics/mplot_1ldn.gif

- the output file of the normal MPlot command is in a form that can be quickly 
converted into an O datablock for those handy with an editor and familiar with 
O datablocks, and could then be used to ramp a model inside O

- you may also want to consider showing how the (main-chain or side-chain) 
torsion angles differ between the structures, e.g. by plotting the circular 
variance of phi and psi - see for instance 
http://xray.bmc.uu.se/cgi-bin/gerard/image_page.pl?image=usf/pics/vmain_1ldn.gif
 - as described here: http://xray.bmc.uu.se/usf/lsqman_man.html#S83 - or a 
multiple-model Ramachandran plot like this 
http://xray.bmc.uu.se/cgi-bin/gerard/image_page.pl?image=usf/pics/mrama_1ldn.gif
 (with the MRama command). The advantage is that no superposition is required 
at all and that any domain movements won't debeautify your results

--dvd




On Tue, 23 Feb 2010, Stephen Graham wrote:

 I am pretty sure you can do this using LSQMAN from Gerard (BluRay?) Kleywegt.
 
 The pertinent commands are MCENTRAL to determine the 'most
 representative structure' (i.e. the one to align upon and show in the
 figure), MALIGN to do the alignment and then MPLOT to calculate a
 'multi-RMSD' for each residue (see manual for details - set the
 'cut-off for printing' to 0 to get all values).
 
 Regards depiction, I think pymol can also represent structures as
 sausages based on their B values:
 cartoon putty
 show cartoon
 
 HTH,
 
 Stephen
 
 On 23 February 2010 01:31, Ethan Merritt merr...@u.washington.edu wrote:
 Hi all,
 
 I am comparing 4 very similar (1.5A rmsd) large (750 residues) structures,
 but struggling to find a way to generate a figure that conveys where they
 are most alike and where they diverge.
 
 Simply drawing a superimposed set of backbone traces results in what looks
 like colored spaghetti.  I don't think that's going to work.
 
 So I had the idea of drawing a single backbone trace, or ribbon diagram,
 and coloring by the RMSD of the four C-alphas at each residue position.
 But I can't find a program that will output this as a table of numbers
 I can use.  All of the multiple structure superposition programs must
 have this information internally.  After all, that's what they are 
 minimizing.
 But do any of the programs provide an option to write it out?
 
 I can get pairwise per-residue deviations by doing SSM superposition in Coot,
 but that doesn't get me to an RMSD for all four structures jointly.
 
Ethan
 
 
 --
 Ethan A Merritt
 Biomolecular Structure Center
 University of Washington, Seattle 98195-7742
 
 
 
 
 -- Dr Stephen Graham
 1851 Research Fellow
 Cambridge Institute for Medical Research
 Wellcome Trust/MRC Building
 Addenbrooke's Hospital, Hills Road
 Cambridge, CB2 0XY, UK
 Phone: +44 1223 762 638
 


Best wishes,

--Gerard

Re: [ccp4bb] smeared spot in diffraction

[MARTYN SYMMONS]

One other thought following from Kelly's point about room temp check is that 
most likely these are thin plates or needles. And I think it was Frank VonD who 
pointed out that some of these thin crystals can be physically stressed by the 
shape of the frozen cryo formed in the cryoloop. So choose loops with care - 
maybe go with litholoops?
 
Martyn
 
Martyn Symmons
MRC-MBU 
Cambridge




- Original Message 
From: Kelly Daughtry kddau...@bu.edu
To: CCP4BB@JISCMAIL.AC.UK
Sent: Thursday, 21 January, 2010 14:27:58
Subject: Re: [ccp4bb] smeared spot in diffraction

Fengxia,
To me the diffraction appears as if the crystals did not freeze well.
So the best option seems to be annealing (as already suggested) or to
try different cryo-protectants.

Did you perform a room-temperature mount? If so, were the spots nice
and round (suggesting the cryo is the cause of the smears)?

I would suggest growing your crystals in the presence of cryo as well.
Glycerol, PEG 400, all the usual suspects.

May I suggest 2-methyl-2,4-pentanediol (MPD). I have had success
adding to well (10 - 20 %) but not in crystal drop, then using 20% as
a cryo (MPD acting to slow diffusion to give better ordered crystals).

Kelly Daughtry

***
Kelly Daughtry
PhD Candidate
Department of Physiology and Biophysics
Boston University School of Medicine
590 Commonwealth Ave
R 390
Boston MA, 02215
(P) 617-358-5548
***



On Thu, Jan 21, 2010 at 9:07 AM, Brad Bennett bradbennet...@gmail.com wrote:
 Hi Fengxia-
 How about increasing the PEG% so you don't have to add as much other
 cryoprotectant?

 Also, have you tried annealing the crystals? I've had success with this when
 I've had samples diffract similarly. The simplest way is by blocking the
 cryostream for 2-3 seconds (repeat 1-2 times) and then shoot. More involved
 way is by actually dismounting your crystals back into cryo buffer for 20-30
 seconds, then remounting and shooting.

 HTH-
 Brad

 On Thu, Jan 21, 2010 at 1:54 AM, Fengxia Liu fengxia@gmail.com wrote:

 Dear all,

 I am trying to diffract one semet-protein, it gave me some clear spots and
 some smeared spots in one diffration, you can find maps attached.
 Mother liquor condition is 0.1MTris-HCL pH8.5+10% PEG4k + 0.2M Li2SO4, I
 have tried mother liquor+25%glycerol, paratone, paratone+ mineral oil, but
 they all gave me such pattern.
 Does anyone know how to solve this? I appreciate any advice.

 Thank you in advance.

 Best wishes,
 Fengxia

Re: [ccp4bb] where I have been going wrong in crystallization?

[MARTYN SYMMONS]

Thanks to those who replied.

The humour comes of course from a slight feeling that there is
some truth in the 'respectful' version. Proteins are wonderful subtle
machines so perhaps we should be careful and thoughtful when we expose them to 
the
horrors of most crystallization conditions!

Which takes me to the original point of my search - which was for a dialysis 
method for lysozyme. I was trying to set up a micro dialysis format for my 
protein but wanted to test it first.  I am using spectrapore 8K dialysis 
membrane and wondered if that would retain lysozyme enough to get crystals (I'd 
like to stick with this cut-off as the physical and sealing properties of other 
membranes might be different). Or if someone has a higher MW cheap alternative 
protein/condition?

Best wishes - agus 'Bliadhna Mhath Ur Dhuibh Uile' as we say in gaelic. 

  Martyn
-
Martyn Symmons
Cambridge

  



- Original Message 
From: David Briggs drdavidcbri...@gmail.com
To: CCP4BB@JISCMAIL.AC.UK
Sent: Monday, 21 December, 2009 17:26:15
Subject: Re: [ccp4bb] where I have been going wrong in crystallization?

Hi Martyn,

this recipe tends to work for me...

Lysozyme: 50 mg/ml in 0.1 M Sodium Acetate pH 4.8
Reagent: 8% w/v Sodium Chloride, 0.1 M Sodium Acetate pH 4.8
Additional Reagents: Index Reagent 8, 22, 28, 31, 34, 40, 58, 59, 69, 86, 88

Mix equal amounts of lysozyme with reagent, incubate at 4 or 22
degrees Celsius. Batch or vapor diffusion works fine.

== Direct copy/paste from http://hamptonresearch.com/experiments.aspx ==

HTH and Happy Christmas.

Cheers,

Dave


David C. Briggs PhD

University of Manchester E-mail:
david.c.bri...@manchester.ac.uk

Twitter: @xtaldave
Skype: DocDCB




2009/12/21 MARTYN SYMMONS martainn_oshioma...@btinternet.com:
 Dear All
  checking out the Lysozyme crystallization methods on the web I liked the 
 Rigaku Instructions that I found:
 (http://www.rigaku.com/protein/crystallization.html)

 ...create a drop of 3ul lysozyme solution, and 3 ul of well solution, 
 respectfully, for a total drop size of 6ul...

 So perhaps sometimes I am just not respectful enough to deserve crystals ?

good wishes to all
   regards,
 Martyn
 ---
 Martyn Symmons
 MRC-MBU Cambridge UK
 'Chan fhiosrach mur feòraich.'
 Gaelic proverb -
  Nothing asked, nothing learned.

[ccp4bb] where I have been going wrong in crystallization?

[MARTYN SYMMONS]

Dear All
  checking out the Lysozyme crystallization methods on the web I liked the 
Rigaku Instructions that I found:
(http://www.rigaku.com/protein/crystallization.html)

...create a drop of 3ul lysozyme solution, and 3 ul of well solution, 
respectfully, for a total drop size of 6ul...

So perhaps sometimes I am just not respectful enough to deserve crystals ?

good wishes to all 
   regards,
 Martyn 
---
Martyn Symmons
MRC-MBU Cambridge UK 
'Chan fhiosrach mur feòraich.'
Gaelic proverb -
  Nothing asked, nothing learned.

Re: [ccp4bb] 3D search for peptide conformers?

[MARTYN SYMMONS]

I think SPASM by Gerard Kleywegt is a great tool for this sort of thing - you 
can set the similarity matrix threshold to make it more or less sequence 
independent (it still calculates the sequence match which is good for looking 
for key residues in the motif like structurally required GLYs). You can also 
restrict it to just CA positions. And you can set the thresholds for the search 
to narrow down on the basis of rmsd.

J Mol Biol. 1999 Jan 29;285(4):1887-97.
Recognition of spatial motifs in protein structures.
Kleywegt GJ.

You can download the program and a search library - currently july 2008 pdb I 
think (culled to 99% seq ident. is the best one). But could be something more 
recent available?

Good luck
Martyn

Martyn Symmons - Cambridge

From: Patrick Loll pat.l...@drexel.edu
To: CCP4BB@JISCMAIL.AC.UK
Sent: Monday, 14 December, 2009 20:02:53
Subject: [ccp4bb] 3D search for peptide conformers?

I have a 10-residue stretch of a protein that adopts an interesting 
conformation; I'd like to know if this conformation occurs in other proteins. 
I'd welcome suggestions for tools that will allow me to to search for this 
peptide conformation in the PDB. 

I naturally thought of DALI, but it seems to require a minimum length of 30 
residues. 

Thanks,

Pat

---
Patrick J. Loll, Ph. D.  
Professor of Biochemistry  Molecular Biology
Director, Biochemistry Graduate Program
Drexel University College of Medicine
Room 10-102 New College Building
245 N. 15th St., Mailstop 497
Philadelphia, PA  19102-1192  USA

(215) 762-7706
pat.l...@drexelmed.edu 

Re: [ccp4bb] Refining residues as rigid bodies

[MARTYN SYMMONS]

Discussing with Fred Vellieux offlist I suggested the following: would it be 
useful to have a sort of 'simulated annealing' of the B-factor values? - in 
refinement I often try resetting them to higher values (Moleman has a function 
for this I think) and then see which ones refine back down nicely - but it 
would be good to have some sort of randomization method - that would be a test 
of how much the values are inherited from the modelling early on - say from the 
MR probe model for example. 

We could do multiple B-factor refinements with different starting kicks to the 
values and look for the best final Rfree or some measure of map quality?

Martyn

Martyn F. Symmons
Cambridge
'Chan fhiosrach mur feòraich.'
Gaelic proverb - Nothing asked, nothing learned.

 






From: Pavel Afonine pafon...@lbl.gov
To: CCP4BB@JISCMAIL.AC.UK
Sent: Wednesday, 2 December, 2009 22:52:45
Subject: Re: [ccp4bb] Refining residues as rigid bodies

Hi,

you can do similar thing (that is resulting in similar outcome) in
phenix.refine by increasing the weight on ADP restraints term. Example:
increase wu or decrease wxu_scale. Although I believe a regular
refinement of individual isotropic ADPs should normally work just fine
at 3A resolution in phenix.refine. 

Pavel.


On 12/1/09 11:18 PM, Frederic VELLIEUX wrote: 
If the problem is B-factor refinement, you can do that easily at low resolution 
with CNS. You just give tight restraints.

You modify the file bindividual.inp

This section:
{* target sigma values for restrained B-factor refinement *}

{* mainchain bonds *}
{===} bsig_main=1.5;
{* mainchain angles *}
{===} asig_main=2.0;

{* sidechain bonds *}
{===} bsig_side=2.0;
{* sidechain angles *}
{===} asig_side=2.5;

I cannot remember in which direction the values have to go. I think up. I have 
done this with a very low resolution structures (4.5 A?) a few years ago, you 
get smoothly varying B values, very tightly restrained. I do not think we 
published that structure, we obtained a higher resolution structure later (I 
don't think the referees would have been very happy seeing B factor refinement 
at low resolution). But it worked.

Fred.

  
Message du 01/12/09 23:51
De : Jason C Porta 
A : CCP4BB@JISCMAIL.AC.UK
Copie à : 
Objet : Re: [ccp4bb] Refining residues as rigid bodies


Basically my reasoning for doing this is a low data-to-parameter ratio, which 
makes B-factor refinement unfeasible. So far I have had nice results with 
breaking the complex into rigid subdomains. So i was basically just thinking 
of a way I could refine the structure best, without using too many parameters.

I see now how this can be done in Phenix. I will give it a try.

Thanks for all of your suggestions!
  

Re: [ccp4bb] Refining residues as rigid bodies

[MARTYN SYMMONS]

IMHO this is a bad idea as the restraints can give undue weight to poorly 
ordered residues in order to favour the tighter distribution that you are 
specifying. 

I have found bindividual.inp to perform at low resolution reasonably well, and 
I think it is more realistic than group B-factors. The overall weighting on 
these B-factor restraints is usually determined automatically by CNS I think.

The target sigmas in the distributed bindividual.inp  file are too tight to 
start with and I would always follow previous suggestions on the bb - mainchain 
bond 2.0, mainchain angle 3.0, sidechain bond 4.5, sidechain angle 6.0. You can 
check the log file afterwards to see if the refinement is actually getting 
close to the values you set. Then you can manually set the B-factor restraint 
weighting if you are not happy with the program's choice. That would be a 
better way to restrain than changing the target distributions?

I think the PHENIX restrained B-factor refinement is very well thought out - 
based on ideas from Ian Tickle's analysis. And I would think that should be 
good at low resolution too - so perhaps worth a try? You can always do just 
B-factor refinement in PHENIX after geometric refinement in CNS if you want.

regards
  Martyn 

Martyn F. Symmons
Cambridge UK
'Chan fhiosrach mur feòraich.'
Gaelic proverb - Nothing asked, nothing learned. 









From: Frederic VELLIEUX frederic.velli...@orange.fr
To: CCP4BB@JISCMAIL.AC.UK
Sent: Wednesday, 2 December, 2009 8:56:01
Subject: Re: [ccp4bb] Refining residues as rigid bodies

Dear Bulletin Board,

I received this information from Axel Bruenger, who rightly corrected me on the 
modification to be made to the bindividual.inp file:

As an fyi, the sigmas should be made smaller to get a narrower B-factor 
differences.

My previous post:

 You modify the file bindividual.inp
 
 This section:
 {* target sigma values for restrained B-factor refinement *}
 
 {* mainchain bonds *}
 {===} bsig_main=1.5;
 {* mainchain angles *}
 {===} asig_main=2.0;
 
 {* sidechain bonds *}
 {===} bsig_side=2.0;
 {* sidechain angles *}
 {===} asig_side=2.5;
 
 I cannot remember in which direction the values have to go. I think up.

So the values have to go down. The precise values I used are somewhere on a DAT 
tape and we have no DAT reader here anymore...

Fred.

Re: [ccp4bb] off topic; alignment keeping upper and lower case

[MARTYN SYMMONS]

Dear All
  - I am looking for a multiple alignment program that will work 
with upper and lower case letters in the sequences and preserve them through to 
the output. I used to use PILEUP but that is no longer supported here. 
   Any suggestions much appreciated.
   regards 
Martyn 

Martyn F. Symmons 
Department of Pathology
University of Cambridge

'Gun fhaillin 'san fhairge..'
(said of Griomasaigh boats)

[ccp4bb] unknown PyMOL script author

[MARTYN SYMMONS]

Dear CCP4ers
Do you recognize your Python code in the script below? I got this from a script 
in the PyMOL wiki last year but now I can't locate it or identify the author. I 
have used this syntax extensively in some structural analysis that I am now 
writing up and I would like to credit the originator (PyMOL and the wiki both 
get a citation obviouslybut I checked and neither Warren Delano nor Jason 
Vertrees is the author). 
 
for a in cmd.index(A//37/CB):
for b in cmd.index(B//LYS/NZ):
cmd.select(s1,%s`%d%a)
cmd.select(s2,%s`%d%b)
if cmd.select((s1|s2) and not ?skip):
if cmd.dist(tmp,s1,s2)  20.0 :
  print '',cmd.iterate(s1|s2,print 
'',segi,resn,resi,name),round(cmd.dist(tmp,s1,s2),3)

Thanks
Best wishes
  Martyn 

Martyn Symmons
Department of Pathology
University of Cambridge

Re: [ccp4bb] Poor electron density - polyAla or PolyGly?

[MARTYN SYMMONS]

Dear Joe
  one thing that has helped me a lot with similar problems is G. 
Kleywegt's SPASM - you make your best guess of the mainchain then add a total 
guess of the sidechains (say with mutate in COOT - choosing the top rotamer). 
Then you cut out the loop and run this in SPASM as a search model looking for 
similar motifs in the pdb. You specify a low cut-off for the substitution 
matrix - to get lots of hits - and use high values (3.0-4.5A) for the initial 
positional errors for the residues - this means you get maybe 100 possible 
structurally similar loops from other proteins. Then you can sort the output on 
BLOSUM similarity score or on RMSD to your initial guess. The BLOSUM search 
criterion is the reason that you add the sidechains. If you get no hits with CA 
and SC (sidechain) then you can switch to CA only.  Or try with a more 
restricted bit of your loop. Or look through the output just for loops that 
share a structurally conserved Gly or Pro with
 your sequence.

If I get any possible hits I then superimpose them  on the electron density to 
see if they give hints how the sidechains might lie and also how the backbone 
H-bonding might run. There are lots of small motifs with conserved sidechain to 
mainchain H-bonding (there is a database of these in Glasgow I think) and so 
you pick up hints from similar loops more often than you might think - from 
completely unrelated proteins. 

I have a script to run SPASM if you want it. 

  Another approach is RAPPER which builds multiple a priori loops very quickly 
and sees if they fit the density. There used to be a RAPPER server... not sure 
if it is still active.
   
Good luck. 
  Martyn 





- Original Message 
From: Joe Smith [EMAIL PROTECTED]
To: CCP4BB@JISCMAIL.AC.UK
Sent: Wednesday, 15 October, 2008 4:11:01 PM
Subject: [ccp4bb] Poor electron density - polyAla or PolyGly?

Hello,
I have been building a protein model (resolution 2.2A) which has one
small loop of 6 residues having poor density. I cannot see any side
chain in this region but I can see relatively poor main-chain density
which at least clearly indicate the loop conformation. I am trying my
best to build polyAla chain in this region. But 3-4 residues end up
coming in disallowed region of Ramachandran plot. I think the problem
is - in this region i can not even build the main chain atoms with
high confidence.
Can I build polyGly in this region? or should I just leave this
region? I just don't feel like leaving this region empty.
Any suggestions in this regards is highly appreciated!
regards
Joe

[ccp4bb] Protein-protein docking

[MARTYN SYMMONS]

I would recommend giving Dave Ritchie's Hex program a go - it is blisteringly 
quick. It's limited to local searches so it is best if you have a reasonable 
idea where you expect the docking surface to be (mutations, NMR shift data 
etc). We used it to refine solutions from searches with crosslinking 
restraints. It works nicely interactively if you explore the receptor surface 
by centring the search on different residues. 

Best wishes
Martyn 

Martyn Symmons
Department of Pathology 
University of Cambridge
  
 

Re: [ccp4bb] oxidised cys

[Martyn Symmons]

A possible modification for cysteine that adds extra density is 
S-(dimethylarsenic) cysteine (CAS). Requires DTT and cacodylate buffer 
conditions however. And does not crosslink so far as I know. 
   
Has been seen in a number of structures from cacodylate conditions - eg. one of 
the Xrcc4 structures 

cheers
  Martyn 

Martyn Symmons
Department of Pathology 
University of Cambridge





 Message Received: Apr 12 2007, 06:06 PM
 From: Flip Hoedemaeker [EMAIL PROTECTED]
 To: [EMAIL PROTECTED]
 Cc: 
 Subject: Re: [ccp4bb] oxidised cys
 
 I've actually seen something like this on disulfides (or at least I think
 so, I havent seen your density obviously), turned out it was model bias in
 MR, if I used a different template for MR the feature went away. This was
 high resolution stuff (~1.0 Å).
 
 Flip
 
 -Original Message-
 From: CCP4 bulletin board [mailto:[EMAIL PROTECTED] On Behalf Of
 [EMAIL PROTECTED]
 Sent: Tuesday, April 10, 2007 20:44
 To: [EMAIL PROTECTED]
 Subject: Re: [ccp4bb] oxidised cys
 
 Hi Stefano,
 
 How certain are you that this link is truly what you think it is? If I
 understand what you're saying - you want to create a (thioperoxythio) link
 - this chemistry should be hideously unstable. Can you explain this using
 disorder, or perhaps the residual density is a symmetry artifact?
 
 Regards,
 
 Artem
 
  Dear all
  in my structure I think I can see an oxidised Cys in cys-SO. Refining
  cys-SO
  I observe a residual density between the oxigen of one oxidised cys and
  the
  one of the other molecule in AU.
  I'd like to try to refine it as cys-SO-OS-cys. I didn't find an example of
  it in the pdb database. Could anyone tell me whether there are other
  cases?
  I guess I just didn't find them.
  Second question:
  How could I explain to refmac that there is the OO bond?
  I tried to write a line similar to the one for SSBOND in the pdb header
  OOBOND   1 CEA A   42CEA D   42
  but refmac couldn't care less...
 
  thanks in advance
 
  Stefano
 
  _
  Express yourself instantly with MSN Messenger! Download today it's FREE!
  http://messenger.msn.click-url.com/go/onm00200471ave/direct/01/
 
 

Re: [ccp4bb] oxidised cys

[Martyn Symmons]

Dear John
 I think this is a thiol-specific reaction - where it has happened to 
Cys residues the Met residues appear normal. I wondered if anyone had ever used 
this on purpose as a heavy atom derivative. Arsenic has quite a good anomalous 
signal too I think. 
  All the best
  Martyn 

Martyn Symmons
Department of Pathology 
University of Cambridge




 Message Received: Apr 13 2007, 01:37 PM
 From: John Walker [EMAIL PROTECTED]
 To: [EMAIL PROTECTED]
 Cc: [EMAIL PROTECTED]
 Subject: Re: [ccp4bb] oxidised cys
 
 Can methionine be modified with these two reagents in a similar manner?
 
 Cheers,
 
 John
 
 -- 
 John R. Walker, Ph.D.
 Structural Genomics Consortium
 University of Toronto
 Toronto, Ontario
 Canada
 
 On 4/13/07, Martyn Symmons [EMAIL PROTECTED] wrote:
  A possible modification for cysteine that adds extra density is 
  S-(dimethylarsenic) cysteine (CAS). Requires DTT and cacodylate buffer 
  conditions however. And does not crosslink so far as I know.
 
  Has been seen in a number of structures from cacodylate conditions - eg. one 
  of the Xrcc4 structures
 
  cheers
Martyn
 
  Martyn Symmons
  Department of Pathology
  University of Cambridge
 
 
 
 
  
   Message Received: Apr 12 2007, 06:06 PM
   From: Flip Hoedemaeker [EMAIL PROTECTED]
   To: [EMAIL PROTECTED]
   Cc:
   Subject: Re: [ccp4bb] oxidised cys
 
   I've actually seen something like this on disulfides (or at least I think
   so, I havent seen your density obviously), turned out it was model bias in
   MR, if I used a different template for MR the feature went away. This was
   high resolution stuff (~1.0 Å).
 
   Flip
 
   -Original Message-
   From: CCP4 bulletin board [mailto:[EMAIL PROTECTED] On Behalf Of
   [EMAIL PROTECTED]
   Sent: Tuesday, April 10, 2007 20:44
   To: [EMAIL PROTECTED]
   Subject: Re: [ccp4bb] oxidised cys
 
   Hi Stefano,
 
   How certain are you that this link is truly what you think it is? If I
   understand what you're saying - you want to create a (thioperoxythio) link
   - this chemistry should be hideously unstable. Can you explain this using
   disorder, or perhaps the residual density is a symmetry artifact?
 
   Regards,
 
   Artem
 
Dear all
in my structure I think I can see an oxidised Cys in cys-SO. Refining
cys-SO
I observe a residual density between the oxigen of one oxidised cys and
the
one of the other molecule in AU.
I'd like to try to refine it as cys-SO-OS-cys. I didn't find an example of
it in the pdb database. Could anyone tell me whether there are other
cases?
I guess I just didn't find them.
Second question:
How could I explain to refmac that there is the OO bond?
I tried to write a line similar to the one for SSBOND in the pdb header
OOBOND   1 CEA A   42CEA D   42
but refmac couldn't care less...
   
thanks in advance
   
Stefano
   
_
Express yourself instantly with MSN Messenger! Download today it's FREE!
http://messenger.msn.click-url.com/go/onm00200471ave/direct/01/