Re: [ccp4bb] The experiment is still very much needed (though AlphaFold helps a lot)

[Debanu Das]

Hi Tom,

Great to see your new analysis and publication. A wonderful addition to
your recent work in this domain/comparisons. Congratulations!

For those trying to keep up with "AI Drug Discovery" (including AlphaFold,
OpenFold, Gen AI, etc) vs "Conventional Drug Discovery" (primarily
crystallography/CryoEM/structural biology/biophysics for this audience), a
few recent papers and news items:

"AI’s potential to accelerate drug discovery needs a reality check"
https://www.nature.com/articles/d41586-023-03172-6

"Sumitomo Schizophrenia Drug Discovered With AI Tech Fails in Two Phase 3
Trials"
https://medcitynews.com/2023/07/sumitomo-schizophrenia-ai-drug-discovery-in-two-clinical-trial/

"AI-created drugs fall off hype cycle"
https://www.beckershospitalreview.com/pharmacy/ai-created-drugs-fall-off-hype-cycle.html

"After years of hype, the first AI-designed drugs fall short in the clinic"
https://endpts.com/first-ai-designed-drugs-fall-short-in-the-clinic-following-years-of-hype/

As a research community, while it is great to see new sci & tech maturing
and adding to the R, it's quite likely that when the dust settles (as
we've seen with many hypes, I think there are ~200 companies of different
sizes and stages claiming to be in the "AI" space, most with undisclosed
and unvalidated methods and approaches), all this could become a strong
*addition* to the drug discovery/structural biology arsenal, *instead of
substituting workhorse solutions and approaches*, potentially *most useful
in cases only when experimental structures are not available or not
immediately possible to kick off the start of projects*.

Best regards,
Debanu
--
Debanu Das,
https://bio.site/debanu_das

On Fri, Dec 1, 2023 at 10:23 AM Mitchell D. Miller <
mitchell.d.mil...@rice.edu> wrote:

> The Baker group has also reported substantial progress in all atom
> predictions
>
> Krishna et al.,
> Generalized Biomolecular Modeling and Design with RoseTTAFold All-Atom
> bioRxiv 2023.10.09.561603
> doi: 10.1101/2023.10.09.561603
> https://www.biorxiv.org/content/10.1101/2023.10.09.561603v1.full
>
> Regards,
> Mitch
>
> Quoting Tom Terwilliger <b6116340b489-dmarc-requ...@jiscmail.ac.uk>:
>
> > Hi Roberto,
> > Thanks for the link to the DeepMind site!  I hadn't seen that and now I
> > read it and the paper that they link to.  It looks to me as though they
> are
> > making significant progress on ligands, dna, rna, but that the program is
> > not yet available, so I don't think it can be tested yet!
> > All the best,
> > Tom T
> >
> >
> > On Fri, Dec 1, 2023 at 5:19 AM Roberto Steiner <
> roberto.stei...@kcl.ac.uk>
> > wrote:
> >
> >> Great paper indeed!
> >>
> >> However quite significant progress seems to have been achieved with
> >> ligands as well (and not only)…
> >>
> >>
> https://urldefense.com/v3/__https://deepmind.google/discover/blog/a-glimpse-of-the-next-generation-of-alphafold/__;!!BuQPrrmRaQ!mWTLfJ8Bjj3YYdp1ZCa8HXyjF-1lq1emf12VPcusoqMN5sh4Eajgq-3UOcaWAERlTNpVWXSdqVPUzhyjIYzPrkIn0P-gtdIjb9GM9FxbyWs$
> >> Has anyone in the community put this to the test?
> >>
> >>
> >> Best wishes
> >> Roberto
> >>
> >>
> >> *Roberto A Steiner*
> >>
> https://urldefense.com/v3/__http://www.steinerlab.org__;!!BuQPrrmRaQ!mWTLfJ8Bjj3YYdp1ZCa8HXyjF-1lq1emf12VPcusoqMN5sh4Eajgq-3UOcaWAERlTNpVWXSdqVPUzhyjIYzPrkIn0P-gtdIjb9GMxbA8Qgg$
> >>
> https://urldefense.com/v3/__https://twitter.com/steiner_lab__;!!BuQPrrmRaQ!mWTLfJ8Bjj3YYdp1ZCa8HXyjF-1lq1emf12VPcusoqMN5sh4Eajgq-3UOcaWAERlTNpVWXSdqVPUzhyjIYzPrkIn0P-gtdIjb9GMZmS2rX0$
> >>
> >> roberto.stei...@kcl.ac.uk
> >> Randall Centre for Cell and Molecular Biophysics
> >> Faculty of Life Sciences and Medicine
> >> King's College London
> >> Room 3.10A
> >> New Hunt's House, Guy's Campus
> >> SE1 1UL, London, UK
> >> Phone 0044 20 78488216
> >> Fax0044 20 78486435
> >>
> >> roberto.stei...@unipd.it
> >> Dipartimento di Scienze Biomediche
> >> Università degli Studi di Padova
> >> Viale G. Colombo 3
> >> 35131 Padova, Italia
> >> Telefono 0039 049 8276409
> >>
> >> *Responses to emails are not expected outside of your normal working
> >> hours.*
> >>
> >>
> >>
> >>
> >>
> >>
> >>
> >>
> >> On 30 Nov 2023, at 22:35, Tom Terwilliger <
> >> b6116340b489-dmarc-requ...@jiscmail.ac.uk> wrote:
> >>
> >> You don't often get email from
> >> b6116340b489-dmarc-re

Re: [ccp4bb] Dry shipper in limbo

[Debanu Das]

Hi Jan, Kevin,

Our experience shipping from San Francisco to CLS has ranged from arrival
on 2nd day to 10th day (both extreme), most commonly on 3rd or 4th day,
irrespective of using the CLS recommended customs broker or FedEx as the
international broker. The main parameter is likely if the package goes
directly to Calgary or is routed via Memphis, and this is dependent on
FedEx operational logistics and not something that can be selected or
planned for.

It's possible that Seattle to Saskatoon is always routed directly, making
it consistently an overnight delivery. Luckily with our 2 local SF Bay Area
synchrotrons that I've been using regularly for about 20 years, we only
ship elsewhere sporadically.

The top of the line dewars in best condition are rated up to around 14 days
I think, and we have personally tested it up to 10 days without any issues,
although typically I would want to restrict them to ~7 days as dry
shippers.

Typically, in my experience, if there are sudden untraceable/no information
status updates on FedEx due to their logistics/missed connection, usually
they show up on the system again in 1-2 days as the package works through
their delayed delivery/shipping process.

Best,
Debanu

On Fri, Jul 28, 2023 at 6:30 PM Jan Abendroth 
wrote:

> Hi all,
>
> In addition to the statements above, our experience has shown that using
> the Customs broker as described on the CLS webpage has been helpful. Our
> weekly shipments from Seattle to Saskatoon typically make it overnight,
> worst case the next day.
> Also, test your dewars. They should keep cold for a good week. With simple
> tests one can identify dewars that are on their way out.
> Glad that your dewar turned up and that the great staff at CLS could fit
> you in.
>
> Cheers,
> Jan
>
> On Fri, Jul 28, 2023 at 1:09 PM Dr. Kevin M Jude 
> wrote:
>
>> Hi all, thanks for the responses on and off list.
>>
>>
>>
>> Yesterday I got my case assigned to someone in the international shipping
>> division, gave a physical description of my shipping case and expressed the
>> perishable nature of the contents. My dewar turned up in Calgary last night
>> and was just delivered to CLS, hopefully in cold condition. My contact at
>> CLS was able to give my time (yesterday) to another user and get me
>> rescheduled for the weekend. I had shipped Monday for Thursday beamtime,
>> hoping to avoid limbo over the weekend, but maybe in the future I’ll aim a
>> little earlier.
>>
>>
>>
>> Besides the on-list responses, I had a couple of helpful off-list replies:
>>
>>
>>
>> BW duct-tapes an airtag into his shipping cases to help keep an eye on
>> them
>>
>>
>>
>> SG says be sure to include USCMA/CUSMA certificate of origin for
>> shipments within North America, along with statement that the samples are
>> non-hazardous, for research purposes, with no commercial value (and don’t
>> assign a high value to the shipment). When contacting FedEx ask for a
>> supervisor in the international shipping division. Customs inspectors may
>> be less tied up late at night.
>>
>>
>>
>>
>>
>> *From: *CCP4 bulletin board  on behalf of Dr.
>> Kevin M Jude 
>> *Date: *Thursday, July 27, 2023 at 8:16 PM
>> *To: *CCP4BB@JISCMAIL.AC.UK 
>> *Subject: *[ccp4bb] Dry shipper in limbo
>>
>> My first adventure in international crystallography is off to an
>> inauspicious start. On Monday, I sent a dry shipper “overnight” from
>> California to Saskatchewan, but it has been stuck in the Memphis FedEx
>> facility for a few days. I’ve gotten several conflicting explanations of
>> the status from FedEx on the phone, but the most likely seems that it has
>> “pre-cleared” customs and yet has not yet made it to Calgary. It’s not
>> clear whether anyone actually knows where the shipping case is, since I was
>> asked to give a physical description of it. Is there anything else I can do
>> from a few thousand miles away, or do I just have to wait this out?
>>
>>
>>
>> --
>>
>> Kevin Jude, PhD
>>
>> Structural Biology Research Specialist, Garcia Lab
>>
>> Howard Hughes Medical Institute
>>
>> Stanford University School of Medicine
>>
>> Beckman B177, 279 Campus Drive, Stanford CA 94305
>>
>> Phone: (650) 723-6431
>>
>>
>> --
>>
>> To unsubscribe from the CCP4BB list, click the following link:
>> https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1
>>
>> --
>>
>> To unsubscribe from the CCP4BB list, click the following link:
>> https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1
>>
>
>
> --
> Jan Abendroth
> UCB BioSciences
> Seattle / Bainbridge Island, WA, USA
> home: jan.abendr...@gmail.com
> work: jan.abendr...@ucb.com
>
> --
>
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1
>



To unsubscribe from the CCP4BB list, click the following link:

Re: [ccp4bb] Ligand superposition!

[Debanu Das]

Hi Rams,

You could consider using LigPlot+, generate LigPlots for all 'n' PDB files,
and then also superimpose all the plots to see the variations in the
protein-ligand interactions of the same ligand bound to different proteins.

Alternatively, you could try to write a program using Coot's lsq ligand;
CCP4MG Superpose equivalent ligand (might take some time to do manually or
few/many at a time but easy to execute and visualize); try with LSQKAB; or
CSD Ligand Overlay (used for ligand-based design when protein structures
are unknown).

Best,
Debanu

On Sat, Jul 15, 2023 at 6:56 AM Boaz Shaanan  wrote:

> Hi Rams,
> It's also worth looking into the Cambridge CSD suit. I'm pretty sure that
> there is a routine there for superimposing small molecules, i.e. ligands
> which will produce the matrices you need. How to put it on a script is
> another question.
> Cheers,
> Boaz
>
> Boaz Shaanan, Ph.D.
> Dept. of Life Sciences
> Ben Gurion University
> Beer Sheva, Israel
>
> On Jul 15, 2023 15:09, "Subramanian, Ramaswamy" 
> wrote:
> Thanks Boaz,
>
> The proteins are all completely different in folds and structures!
>
>
> Rams
> subra...@purdue.edu
>
>
>
> On Jul 15, 2023, at 8:04 AM, Boaz Shaanan  wrote:
>
>  *External Email*: Use caution with attachments, links, or sharing
> data 
> Hi,
> -You may want to send a query to the UCSF chimerax/chimera people. They
> may suggest how to this using scripts.
> - I have not done this on 50 structures, only on few ones. In my
> experience, if the proteins are sufficiently similar there is a good chance
> that superimposing the proteins will also bring the ligands to near
> superposition.
> -Using chimera/chimerax you can select protein residues within certain
> radius from the ligands, write out the pdb's and then perhaps superimpose
> those smaller pdb's. I have not tried this.
> Sorry I can't be of more help.
> Cheers,
> Boaz
>
>
>
>
>
>
>
>
>
>
>
> *Boaz Shaanan, Ph.D.  Dept. of
> Life Sciences   Ben-Gurion University
> of the Negev   Beer-Sheva
> 84105
> Israel
>
> E-mail: bshaa...@bgu.ac.il  Phone: 972-8-647-2220
>Fax:   972-8-647-2992 or 972-8-646-1710*
>
>
>
>
>
>
> --
> *From:* CCP4 bulletin board on behalf of Subramanian, Ramaswamy
> *Sent:* Friday, July 14, 2023 10:49 PM
> *To:* CCP4BB@JISCMAIL.AC.UK
> *Subject:* [ccp4bb] Ligand superposition!
>
> Hi All,
>
> I have over 50 pdb files that I have downloaded from PDB that all have the
> same ligand bound.
>
> I wamt to superpose the ligands (and move the protein coordinates using
> that matrix).  The goal is for me to see how difference in ligand
> environments in different complexes.
>
> Is there an easy way to do it?  I am sure it has been done before and I do
> not want to reinvent the wheel.
>
> Thanks.
>
>
>
> Rams
> subra...@purdue.edu
>
>
>
>
> --
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1
>
> --
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1
>
>
>
> --
>
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1
>



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1

This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing list 
hosted by www.jiscmail.ac.uk, terms & conditions are available at 
https://www.jiscmail.ac.uk/policyandsecurity/

Re: [ccp4bb] To Trim or Not to To Trim

[Debanu Das]

Yes, zero occupancy would reflect that. But not the coordinates in any
proper way. Then what would be the point of associating occupancy and
B-factors with totally incorrect coordinates?
Debanu

On Fri, Mar 10, 2023 at 8:58 AM Jurgen Bosch  wrote:

> Going back to RIP phasing methods :-)
> So Harry in your particular case occupancy of zero would actually reflect
> reality for those “combusted” atoms.
>
> Jürgen
>
> > On Mar 10, 2023, at 11:56 AM, Harry Powell <
> 193323b1e616-dmarc-requ...@jiscmail.ac.uk> wrote:
> >
> > Hi folks
> >
> > One other thing that I haven’t noticed anyone mentioning yet (sorry to
> those who have mentioned it!!) is that you may not see your sidechain atoms
> in density because they are not there at all, in spite of what you may have
> had in the original protein, or even if the atoms were really there in the
> crystal _before_ exposure to the beam.
> >
> > The coordinates are supposed to be what you actually find, not what you
> hope is there.
> >
> > Just my two ha’porth
> >
> > Harry
> >
> > 
> >
> > To unsubscribe from the CCP4BB list, click the following link:
> > https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1
> >
> > This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a
> mailing list hosted by www.jiscmail.ac.uk, terms & conditions are
> available at https://www.jiscmail.ac.uk/policyandsecurity/
>
> 
>
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1
>
> This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a
> mailing list hosted by www.jiscmail.ac.uk, terms & conditions are
> available at https://www.jiscmail.ac.uk/policyandsecurity/
>
-- 
---
LinkedIn: www.linkedin.com/in/debanudas
Cal Alumni: cal.berkeley.edu/debanudas



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1

This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing list 
hosted by www.jiscmail.ac.uk, terms & conditions are available at 
https://www.jiscmail.ac.uk/policyandsecurity/

Re: [ccp4bb] Structural alignment and classification

[Debanu Das]

Try POSA and FATCAT from Godzik group for structural clusters/trees and
multiple/flexible structure alignments:

https://academic.oup.com/nar/article/48/W1/W60/5848494

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4086100/

https://posa.godziklab.org/
https://fatcat.godziklab.org/

Best regards,
Debanu

On Thu, Mar 9, 2023 at 3:57 PM Jon Cooper <
488a26d62010-dmarc-requ...@jiscmail.ac.uk> wrote:

> Coot's ssm superposition does give the C-alpha rmsd, though, most easily
> seen if you run it from a terminal window? I guess it can't be miles
> different from a least-squares fit ;-?
>
>
> Best wishes, Jon Cooper. jon.b.coo...@protonmail.com
>
> Sent from Proton Mail mobile
>
>
>
>  Original Message 
> On 9 Mar 2023, 13:55, Paul Emsley < pems...@mrc-lmb.cam.ac.uk> wrote:
>
>
> On 06/03/2023 07:35, Armando Albert wrote: > I want to align several
> structures we obtained from a fragment screening campaign and cluster them
> according to RMSD. > To what end, may I ask? I would use LSQKAB, parse the
> log files for the RMSD and create an input file for R. (One cannot just do
> this in Coot, because, for LSQ fitting, Coot only returns the
> transformation matrix, not any fitting statistics - in retrospect this is
> obviously an oversight.) Paul.
>  To
> unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1 This
> message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing
> list hosted by www.jiscmail.ac.uk, terms & conditions are available at
> https://www.jiscmail.ac.uk/policyandsecurity/
> --
>
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1
>
> --
---
LinkedIn: www.linkedin.com/in/debanudas
Cal Alumni: cal.berkeley.edu/debanudas



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1

This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing list 
hosted by www.jiscmail.ac.uk, terms & conditions are available at 
https://www.jiscmail.ac.uk/policyandsecurity/

Re: [ccp4bb] To Trim or Not to To Trim

[Debanu Das]

In fact, for non-SBs using the PDB, it is far likelier that they will
recognize truncated side chains and thus seek help or know what to do
themselves, compared to recognizing incorrectly modeled rotamers (as they
won’t be checking electron density likely) and atoms with zero occupancy
and B-factors, which could lead to fairly incorrect biological
interpretations.
Best,
Debanu

On Thu, Mar 9, 2023 at 7:55 PM Debanu Das  wrote:

> We dealt with this in-depth during structural genomics days when we
> deposited over 1500 novel, high-quality, experimentally-phased structures
> into the PDB. Think it’s prudent to trim/truncate side chains without
> reliable density.
>
> Non-structural biologists using PDB structures without expert help can err
> in any of these scenarios: misinterpreting most common/random rotamer, zero
> occupancy atoms, B-factors, etc.
>
> What is the value of populating the PDB, which is a structural model
> repository, with such information that is not there, i.e., reliable
> structural model?
>
> Any trained crystallographer/structural biologist can easily add in side
> chain information if needed for modeling/computational chemistry reasons.
>
> Best regards,
> Debanu
>
> On Thu, Mar 9, 2023 at 6:50 PM Jurgen Bosch  wrote:
>
>> I’d say no trimming to side chains for the following reason: There are
>> non-structural biologists using PDB files and if atoms are missing they
>> don’t know what to do. A better approach is where no side chain density
>> allows support of placement, pick the most common rotamer and set the
>> occupancy to zero for those atoms lacking density support. More work for
>> you but more accurate in my opinion.
>>
>> Jürgen
>>
>>
>> ___
>> Jürgen Bosch, PhD, MBA
>> Center for Global Health & Diseases
>> Case Western Reserve University
>> Cleveland, OH 44106
>> https://www.linkedin.com/in/jubosch/
>>
>> CEO & Co-Founder at InterRayBio, LLC
>>
>>
>>
>> On Mar 9, 2023, at 9:45 PM, Bernhard Lechtenberg <
>> 968307750321-dmarc-requ...@jiscmail.ac.uk> wrote:
>>
>> Hi Rhys,
>>
>> I am also all for leaving side chains and letting the B-factors deal with
>> the weak/absent density.
>>
>> I don’t think there is a consensus, but I kind of remember that somebody
>> did a poll a few years ago and if I remember correctly the main approaches
>> were the one described above, or trimming the side-chains.
>>
>> Bernhard
>>
>> *Bernhard C. Lechtenberg* PhD
>> NHMRC Emerging Leadership Fellow
>> Laboratory Head
>> Ubiquitin Signalling Division​​
>> E lechtenber...@wehi.edu.au
>> T +61 3 9345 2217
>>
>>
>>
>> *From: *CCP4 bulletin board  on behalf of Rhys
>> Grinter <22087c81e8c6-dmarc-requ...@jiscmail.ac.uk>
>> *Date: *Friday, 10 March 2023 at 12:26 pm
>> *To: *CCP4BB@JISCMAIL.AC.UK 
>> *Subject: *[ccp4bb] To Trim or Not to To Trim
>> Hi All,
>>
>> I'm trying to crowdsource an opinion on how people deal with modelling
>> side chains with poorly resolved electron or cryoEM density.
>>
>> My preference is to model the sidechain and allow the B-factors to go
>> high in refinement to represent that the side chain is flexible. However,
>> I'm aware that some people truncate sidechains if density is not present to
>> justify modelling. I've also seen models where the sidechain is modelled
>> but with zero occupancy if density isn't present.
>>
>> Is there a consensus and justifying arguments for why one approach is
>> better?
>>
>> Cheers,
>>
>> Rhys
>>
>>
>> --
>>
>> To unsubscribe from the CCP4BB list, click the following link:
>> https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1
>>
>> --
>>
>> To unsubscribe from the CCP4BB list, click the following link:
>> https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1
>>
>>
>>
>> --
>>
>> To unsubscribe from the CCP4BB list, click the following link:
>> https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1
>>
> --
> ---
> LinkedIn: www.linkedin.com/in/debanudas
> Cal Alumni: cal.berkeley.edu/debanudas
>
-- 
---
LinkedIn: www.linkedin.com/in/debanudas
Cal Alumni: cal.berkeley.edu/debanudas



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1

This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing list 
hosted by www.jiscmail.ac.uk, terms & conditions are available at 
https://www.jiscmail.ac.uk/policyandsecurity/

Re: [ccp4bb] To Trim or Not to To Trim

[Debanu Das]

We dealt with this in-depth during structural genomics days when we
deposited over 1500 novel, high-quality, experimentally-phased structures
into the PDB. Think it’s prudent to trim/truncate side chains without
reliable density.

Non-structural biologists using PDB structures without expert help can err
in any of these scenarios: misinterpreting most common/random rotamer, zero
occupancy atoms, B-factors, etc.

What is the value of populating the PDB, which is a structural model
repository, with such information that is not there, i.e., reliable
structural model?

Any trained crystallographer/structural biologist can easily add in side
chain information if needed for modeling/computational chemistry reasons.

Best regards,
Debanu

On Thu, Mar 9, 2023 at 6:50 PM Jurgen Bosch  wrote:

> I’d say no trimming to side chains for the following reason: There are
> non-structural biologists using PDB files and if atoms are missing they
> don’t know what to do. A better approach is where no side chain density
> allows support of placement, pick the most common rotamer and set the
> occupancy to zero for those atoms lacking density support. More work for
> you but more accurate in my opinion.
>
> Jürgen
>
>
> ___
> Jürgen Bosch, PhD, MBA
> Center for Global Health & Diseases
> Case Western Reserve University
> Cleveland, OH 44106
> https://www.linkedin.com/in/jubosch/
>
> CEO & Co-Founder at InterRayBio, LLC
>
>
>
> On Mar 9, 2023, at 9:45 PM, Bernhard Lechtenberg <
> 968307750321-dmarc-requ...@jiscmail.ac.uk> wrote:
>
> Hi Rhys,
>
> I am also all for leaving side chains and letting the B-factors deal with
> the weak/absent density.
>
> I don’t think there is a consensus, but I kind of remember that somebody
> did a poll a few years ago and if I remember correctly the main approaches
> were the one described above, or trimming the side-chains.
>
> Bernhard
>
> *Bernhard C. Lechtenberg* PhD
> NHMRC Emerging Leadership Fellow
> Laboratory Head
> Ubiquitin Signalling Division​​
> E lechtenber...@wehi.edu.au
> T +61 3 9345 2217
>
>
>
> *From: *CCP4 bulletin board  on behalf of Rhys
> Grinter <22087c81e8c6-dmarc-requ...@jiscmail.ac.uk>
> *Date: *Friday, 10 March 2023 at 12:26 pm
> *To: *CCP4BB@JISCMAIL.AC.UK 
> *Subject: *[ccp4bb] To Trim or Not to To Trim
> Hi All,
>
> I'm trying to crowdsource an opinion on how people deal with modelling
> side chains with poorly resolved electron or cryoEM density.
>
> My preference is to model the sidechain and allow the B-factors to go high
> in refinement to represent that the side chain is flexible. However, I'm
> aware that some people truncate sidechains if density is not present to
> justify modelling. I've also seen models where the sidechain is modelled
> but with zero occupancy if density isn't present.
>
> Is there a consensus and justifying arguments for why one approach is
> better?
>
> Cheers,
>
> Rhys
>
>
> --
>
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1
>
> --
>
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1
>
>
>
> --
>
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1
>
-- 
---
LinkedIn: www.linkedin.com/in/debanudas
Cal Alumni: cal.berkeley.edu/debanudas



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1

This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing list 
hosted by www.jiscmail.ac.uk, terms & conditions are available at 
https://www.jiscmail.ac.uk/policyandsecurity/

Re: [ccp4bb] Future Diffraction Methods

[Debanu Das]

Consider merging with the Annual ACA meeting. The ACA meeting can also
benefit from more X-ray diffraction methods rigor and training and it will
help to elevate the continuity and history of the ACA and its previous and
current member participation and contributions in the field.
Best,
Debanu

On Tue, Dec 20, 2022 at 6:40 PM Orville, Allen (DLSLtd,RAL,LSCI) <
66b6f959eb28-dmarc-requ...@jiscmail.ac.uk> wrote:

> Hi,
>
>
>
> Thanks James, for organising our last GRC meeting.
>
>
>
> In the past few years, I’ve also attended one or more Annual BioXFEL
> International Conferences (open, but ending?) and Ringberg Workshop on
> Science with FELs (by invitation only). Both were excellent meetings held
> in the winter, complemented the GRC in diffraction methods, and benefited
> from relatively small attendance with very focused discussions.  I will
> also add that dynamic structural biology is a rapidly growing area that can
> include simultaneous functional, spectroscopic, and structural data
> collection from the same samples, and using synchrotron, XFEL, and/or
> electron sources. Moreover, many of the other life-science centric GRC’s
> also include significant structural data in their frontier programs and
> discussions (e.g. metals in biology, hemes and tetrapyrroles, enzymes,
> etc.). We are fundamental to biochemistry R… a broad community indeed.
>
>
>
> AMO
>
> ~ ~ ~ ~ ~
>
> Allen M. Orville, Ph.D.
>
> Wellcome Investigator and Royal Society Wolfson Fellow
>
> Principal Scientist, XFEL Hub at Diamond Light Source
>
> Harwell Science and Innovation Campus
>
> Didcot, Oxfordshire
>
> OX11 0DE
>
> United Kingdom
>
>
>
> Phone: +44 (0) 1235 567505
>
> Mobile: +44 (0) 7471 026061
>
> email: allen.orvi...@diamond.ac.uk
>
>
>
> AMO site:
> https://www.diamond.ac.uk/Instruments/Mx/XFEL-Hub/Staff/Orville.html
>
> XFEL-Hub site: https://www.diamond.ac.uk/Instruments/Mx/XFEL-Hub.html
>
>
>
>
>
>
>
>
>
>
>
> *From: *CCP4 bulletin board  on behalf of Frank
> von Delft 
> *Date: *Monday, 19 December 2022 at 09:22
> *To: *CCP4BB@JISCMAIL.AC.UK 
> *Subject: *Re: [ccp4bb] Future Diffraction Methods
>
> Shoot... that sucks.
>
> Yes, we do need something!
>
> "Structural Biology Methods"?
>
>
> More interesting question:  does it need the GRC to host?  What about
> reimagining the CCP4 study weekend - the question keeps coming up there
> anyway.
>
> That's one for CCP4 WG1 to discuss - they're meeting straight after New
> Year, so maybe James, you and Ivo should hop on a zoom and kick the idea
> around.
>
> Frank
>
>
>
> On 16/12/2022 22:10, James Holton wrote:
> > I want to thank everyone who attended the 2022 Gordon Research
> > Conference and Gordon Research Seminar on Diffraction Methods in
> > Structural Biology, as well as all those who contributed to these
> > great gatherings in the past.  It was an outstanding meeting if I do
> > say so myself. Not just because it had been so long without in-person
> > interaction, not just because we had zero covid cases (which I see as
> > no small feat of Mind over Virus), but because of this amazing
> > community. It is rare in this world to have such a strong spirit of
> > collaboration, camaraderie and openness in undertakings as high-impact
> > as this. Surmounting the barriers to atomic-detail imaging of
> > biological systems has never been more exciting and more relevant.  I
> > am proud to be a part of it, and honored to have served as Chair.
> >
> > It is therefore with heavy heart that I report to this community that
> > I was the last Chair of the Diffraction Methods GRC.
> >
> > The GRC Conference Evaluation Committee
> > (https://www.grc.org/about/conference-evaluation-committee/) voted
> > this year to discontinue the Diffraction Methods GRC and GRS. This
> > ends a 46-year tradition that I feel played a vital, and vibrant role
> > in the work of the people who answer questions on this BB. The reason
> > given was insufficient attendance.  All other metrics, such as
> > evaluation surveys and demographics were very strong. I have tried to
> > appeal, but I'm told the vote was unanimous and final. I understand
> > that like so many conference organizing bodies the GRC is having to
> > make tough financial decisions. I must say I disagree with this one,
> > but it was not my decision to make.
> >
> > Many of the past and elected Chairs have been gathering and discussing
> > how to replace the Diffraction Methods GRC/GRS going forward. Many
> > great ideas, advice and perspectives have been provided, but that is a
> > select group. I feel it is now time to open up this discussion to the
> > broader community of structural methods developers and practitioners.
> > There are some important questions to ask:
> >
> > * How do we define this community?
> > Yes, many of us do cryoEM too, but is that one methods
> > meeting? or two?
> > * Does this community need a new diffraction methods meeting?
> > As in one meeting or zero?
> > * Should we merge 

Re: [ccp4bb] Another folding AI

[Debanu Das]

Amen.

On Thu, Nov 3, 2022 at 5:19 AM Frank von Delft 
wrote:

> Great, now we have two random number generators.
>
>
> On 03/11/2022 12:10, David J. Schuller wrote:
>
> https://www.biorxiv.org/content/10.1101/2022.07.20.500902v2
>
> Evolutionary-scale prediction of atomic level protein structure with a
> language model
> 
> doi: https://doi.org/10.1101/2022.07.20.500902
>
> Abstract
>
> Artificial intelligence has the potential to open insight into the
> structure of proteins at the scale of evolution. It has only recently been
> possible to extend protein structure prediction to two hundred million
> cataloged proteins. Characterizing the structures of the exponentially
> growing billions of protein sequences revealed by large scale gene
> sequencing experiments would necessitate a breakthrough in the speed of
> folding. Here we show that direct inference of structure from primary
> sequence using a large language model enables an order of magnitude
> speed-up in high resolution structure prediction. Leveraging the insight
> that language models learn evolutionary patterns across millions of
> sequences, we train models up to 15B parameters, the largest language model
> of proteins to date. As the language models are scaled they learn
> information that enables prediction of the three-dimensional structure of a
> protein at the resolution of individual atoms. This results in prediction
> that is up to 60x faster than state-of-the-art while maintaining resolution
> and accuracy. Building on this, we present the ESM Metagenomic Atlas. This
> is the first large-scale structural characterization of metagenomic
> proteins, with more than 617 million structures. The atlas reveals more
> than 225 million high confidence predictions, including millions whose
> structures are novel in comparison with experimentally determined
> structures, giving an unprecedented view into the vast breadth and
> diversity of the structures of some of the least understood proteins on
> earth.
> Competing Interest Statement
>
> The authors have declared no competing interest.
>
>
> ===
>  All Things Serve the Beam
>  ===
>  David J. Schuller
>  modern man in a post-modern world
>  MacCHESS, Cornell University
>  schul...@cornell.edu
>
> --
>
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1
>
>



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1

This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing list 
hosted by www.jiscmail.ac.uk, terms & conditions are available at 
https://www.jiscmail.ac.uk/policyandsecurity/

Re: [ccp4bb] open review?

[Debanu Das]

Hi Tim,
"The only acceptable channel for communication about the review of an NIH
grant application after submission" quote from its section refers
specifically only to how the PI/authorized organized representative can
contact the NIH, i.e., they cannot directly communicate with reviewers and
so on.
It does not bypass, and is not related to the other aspects of the larger
quote I had or the entire set of rules in that link...:)
Cheers,
Debanu

On Tue, Jun 28, 2022 at 2:26 AM Tim Gruene  wrote:

> Hi Debanu,
>
> the section of your quote of the NIH rules is introduced with
> "The only acceptable channel for communication about the review of an
> NIH grant application after submission ", i.e. it would not restrict
> James to upload the draft before(!) he submits. James might want to
> double-check with the NIH, though ;-)
>
> Cheers,
> Tim
>
>
> On Mon, 27 Jun 2022 13:31:35
> -0700 Debanu Das  wrote:
>
> > Hi James,
> >
> > I think it is quite different for publications/open publications
> > (investigator initiated submissions on outcomes of grant-funded
> > research, which are meant for public dissemination, even though all
> > patentable IP is still employed-owned, whether a university, a
> > national lab or a company) vs grant applications (which typically go
> > through university sponsored research or grants admin and there are
> > rules around funding agreements and ownership between universities
> > and funding/sponsoring agencies and NDA/confidentiality agreements
> > between funding agencies and reviewers).
> >
> > I doubt that you can freely upload grant applications (to biorxiv or
> > any similar portal) and reviews but you can check with the
> > LBL/Berkeley (or funding agency) office on that. I think confidential
> > sharing of your application and reviews with collaborators or other
> > third parties that include NDAs is allowed.
> >
> > An excerpt below but more details also available at this link (which
> > also pertains to the larger issue being discussed and which I alluded
> > to earlier):
> >
> > From:
> > https://grants.nih.gov/grants/guide/notice-files/NOT-OD-22-044.html
> > "Maintaining security and confidentiality in the NIH peer review
> > process is essential for safeguarding the exchange of scientific
> > opinions and evaluations without fear of reprisal; protecting trade
> > secrets or other proprietary, sensitive and/or confidential
> > information; providing reliable input to the agency about research
> > projects to support; and safeguarding the NIH research enterprise
> > against the misappropriation of research and development to the
> > detriment of national or economic security. In addition, maintaining
> > integrity in the peer review process is important for maintaining
> > public trust in science.
> >
> > This Notice reminds all participants and stakeholders in the NIH peer
> > review process of federal statutes, regulations, and NIH policies
> > regarding peer review security and confidentiality; their
> > responsibilities for abiding by those rules; and possible actions
> > that the NIH (in coordination with other offices) may take and
> > consequences that may ensue from a violation of those rules.
> > Participants and stakeholders include but are not limited to"
> >
> > Best regards,
> >
> > Debanu
> >
> > On Mon, Jun 27, 2022 at 1:13 PM James Holton  wrote:
> >
> > > Hey Debanu,
> > >
> > > Hmm. Last time I did it I didn't have to go through any IP lawyers
> > > to upload a pre-print to biorxiv.  What I was thinking of is
> > > something similar to that.  Researchers, on their own, deciding to
> > > upload their applications and reviews.  What would be the
> > > motivation? Well, I imagine it is not an uncommon situation where
> > > you might want help from more than just the reviewers on how to
> > > revise your application. I know I always try to get all the help I
> > > can get.
> > >
> > > Might even be able to use biorxiv to do it?  Or am I missing
> > > something?
> > >
> > > -James Holton
> > > MAD Scientist
> > >
> > >
> > > On 6/27/2022 12:16 PM, Debanu Das wrote:
> > >
> > > Thinking about it some more, I think all the materials (patentable
> > > IP or trade secrets, which in the US are IP and under Defense of
> > > Trade Secrets Act) of a researcher are owned by the university. So
> > > just getting across tech transfer/IP of individual univs would be a
> > >

Re: [ccp4bb] open review?

[Debanu Das]

Hi James,

I think it is quite different for publications/open publications
(investigator initiated submissions on outcomes of grant-funded research,
which are meant for public dissemination, even though all patentable IP is
still employed-owned, whether a university, a national lab or a company) vs
grant applications (which typically go through university sponsored
research or grants admin and there are rules around funding agreements and
ownership between universities and funding/sponsoring agencies and
NDA/confidentiality agreements between funding agencies and reviewers).

I doubt that you can freely upload grant applications (to biorxiv or any
similar portal) and reviews but you can check with the LBL/Berkeley (or
funding agency) office on that. I think confidential sharing of your
application and reviews with collaborators or other third parties that
include NDAs is allowed.

An excerpt below but more details also available at this link (which also
pertains to the larger issue being discussed and which I alluded to
earlier):

From: https://grants.nih.gov/grants/guide/notice-files/NOT-OD-22-044.html
"Maintaining security and confidentiality in the NIH peer review process is
essential for safeguarding the exchange of scientific opinions and
evaluations without fear of reprisal; protecting trade secrets or other
proprietary, sensitive and/or confidential information; providing reliable
input to the agency about research projects to support; and safeguarding
the NIH research enterprise against the misappropriation of research and
development to the detriment of national or economic security. In addition,
maintaining integrity in the peer review process is important for
maintaining public trust in science.

This Notice reminds all participants and stakeholders in the NIH peer
review process of federal statutes, regulations, and NIH policies regarding
peer review security and confidentiality; their responsibilities for
abiding by those rules; and possible actions that the NIH (in coordination
with other offices) may take and consequences that may ensue from a
violation of those rules. Participants and stakeholders include but are not
limited to"

Best regards,

Debanu

On Mon, Jun 27, 2022 at 1:13 PM James Holton  wrote:

> Hey Debanu,
>
> Hmm. Last time I did it I didn't have to go through any IP lawyers to
> upload a pre-print to biorxiv.  What I was thinking of is something similar
> to that.  Researchers, on their own, deciding to upload their applications
> and reviews.  What would be the motivation? Well, I imagine it is not an
> uncommon situation where you might want help from more than just the
> reviewers on how to revise your application. I know I always try to get all
> the help I can get.
>
> Might even be able to use biorxiv to do it?  Or am I missing something?
>
> -James Holton
> MAD Scientist
>
>
> On 6/27/2022 12:16 PM, Debanu Das wrote:
>
> Thinking about it some more, I think all the materials (patentable IP or
> trade secrets, which in the US are IP and under Defense of Trade Secrets
> Act) of a researcher are owned by the university. So just getting across
> tech transfer/IP of individual univs would be a massive hurdle before
> thinking of being able to upload grants proposals for sharing.
>
> And funding agencies would first also have to negotiate (and convince)
> with all univs to allow it, even if somehow taxpayers and funding agencies
> could be first convinced about the need or value in doing this. In fact, in
> that scenario, there would actually be no need for a new system to share
> proposals. All funding agencies just have to open up a portal to access
> submitted grants (and I'm quite sure the agencies already have massive
> security around hacking attempts to access all this material).
>
> Cheers,
> Debanu
>
> On Mon, Jun 27, 2022 at 11:58 AM Debanu Das  wrote:
>
>> Dear John,
>>
>> For sure it is an aspiration as a society and as a civilization: to think
>> beyond individual nations. And for that we have some examples as you
>> mentioned at the scientific (IUCr, PDB) and political level (UN). We also
>> have the EU, ASEAN, NATO, etc.
>>
>> However, despite having these organizations, I think even within most of
>> them, for critical strategic information that dictates competitiveness and
>> preparation, sharing is restricted to within the group (at least for the
>> political ones). For that matter, even individual agencies within countries
>> often have restrictions in data and materials sharing.
>>
>> I think if we solve the issue of national competitiveness, social
>> inequality, etc first, we will not even have to discuss if there could be
>> issues openly and globally sharing grant proposals. I guess the counter
>> proposal could be made that may

Re: [ccp4bb] open review?

[Debanu Das]

Thinking about it some more, I think all the materials (patentable IP or
trade secrets, which in the US are IP and under Defense of Trade Secrets
Act) of a researcher are owned by the university. So just getting across
tech transfer/IP of individual univs would be a massive hurdle before
thinking of being able to upload grants proposals for sharing.

And funding agencies would first also have to negotiate (and convince) with
all univs to allow it, even if somehow taxpayers and funding agencies could
be first convinced about the need or value in doing this. In fact, in that
scenario, there would actually be no need for a new system to share
proposals. All funding agencies just have to open up a portal to access
submitted grants (and I'm quite sure the agencies already have massive
security around hacking attempts to access all this material).

Cheers,
Debanu

On Mon, Jun 27, 2022 at 11:58 AM Debanu Das  wrote:

> Dear John,
>
> For sure it is an aspiration as a society and as a civilization: to think
> beyond individual nations. And for that we have some examples as you
> mentioned at the scientific (IUCr, PDB) and political level (UN). We also
> have the EU, ASEAN, NATO, etc.
>
> However, despite having these organizations, I think even within most of
> them, for critical strategic information that dictates competitiveness and
> preparation, sharing is restricted to within the group (at least for the
> political ones). For that matter, even individual agencies within countries
> often have restrictions in data and materials sharing.
>
> I think if we solve the issue of national competitiveness, social
> inequality, etc first, we will not even have to discuss if there could be
> issues openly and globally sharing grant proposals. I guess the counter
> proposal could be made that maybe more sharing of more information will
> eventually lead to equity everywhere (which to some extent is reflected in
> the open sharing of publications).
>
> But for now, I think there are practicality hurdles to cross on these,
> which is why I mentioned "workable" in my initial response. Just in the
> last few years, we have seen examples of more and more focus on IP theft,
> computer hacking to steal research data from organizations and companies,
> more focus on ensuring confidentiality of the peer review process, and
> computer security to avoid leaks of material, and so on.
>
> Not trying to be cynical here, I think it is great for us as a community
> to always have an eye on a larger and nobler purpose while working within
> current practicalities and frameworks.
>
> Thank you.
>
> Best regards,
> Debanu
>
> On Mon, Jun 27, 2022 at 11:18 AM John R Helliwell 
> wrote:
>
>> Dear Debanu,
>> There is indeed much at stake here.
>> Would I do it now, share my proposals, No.
>> Would I do it if funders’ rules required it. Yes.
>> When might funders’ rules require it eg when Tax payers insist that the
>> priority is achieving societal goals asap. Might that happen in the
>> foreseeable future? I don’t think so because we are as scientists good at
>> thinking so far out of the box, such as the internet, or from the 19the
>> century electricity and magnetism, the tax payer sees the benefit of an
>> individual’s curiosity driven research.
>> The bigger point is can we also think beyond individual nations?
>> We know we can: the UN, International Council for Science, IUCr……
>> So, it probably isn’t a one size fits all idea that James has put forward…
>> Best wishes,
>> John
>>
>>
>>
>> Emeritus Professor John R Helliwell DSc
>>
>>
>>
>>
>> On 27 Jun 2022, at 19:03, Debanu Das  wrote:
>>
>> 
>> >So, 2nd question is: would you do it? Would you upload your application
>> >into the public domain for all to see? What about the reviewer comments?
>> >If not, why not?  Afraid people will steal your ideas? Well, once
>> >something is public, its pretty clear who got the idea first.
>>
>> I do not think this ("upload your application into the public domain for
>> all to see") is a workable or desirable idea for a variety of reasons.
>> There are far greater issues that just about getting credit for your ideas.
>> Which is somewhat of an academic and personal pursuit.
>>
>> For one, the entire R paradigm and programs and IP of entire nations
>> (which seems primarily would be the US and potentially some EU countries
>> under this case who if at all choose to sign up for this), universities,
>> companies (business grants) and funding agencies will wreak havoc (~30-40%
>> of US GDP). We already know there is a lopsided distribution of which
>&g

Re: [ccp4bb] open review?

[Debanu Das]

Dear John,

For sure it is an aspiration as a society and as a civilization: to think
beyond individual nations. And for that we have some examples as you
mentioned at the scientific (IUCr, PDB) and political level (UN). We also
have the EU, ASEAN, NATO, etc.

However, despite having these organizations, I think even within most of
them, for critical strategic information that dictates competitiveness and
preparation, sharing is restricted to within the group (at least for the
political ones). For that matter, even individual agencies within countries
often have restrictions in data and materials sharing.

I think if we solve the issue of national competitiveness, social
inequality, etc first, we will not even have to discuss if there could be
issues openly and globally sharing grant proposals. I guess the counter
proposal could be made that maybe more sharing of more information will
eventually lead to equity everywhere (which to some extent is reflected in
the open sharing of publications).

But for now, I think there are practicality hurdles to cross on these,
which is why I mentioned "workable" in my initial response. Just in the
last few years, we have seen examples of more and more focus on IP theft,
computer hacking to steal research data from organizations and companies,
more focus on ensuring confidentiality of the peer review process, and
computer security to avoid leaks of material, and so on.

Not trying to be cynical here, I think it is great for us as a community to
always have an eye on a larger and nobler purpose while working within
current practicalities and frameworks.

Thank you.

Best regards,
Debanu

On Mon, Jun 27, 2022 at 11:18 AM John R Helliwell 
wrote:

> Dear Debanu,
> There is indeed much at stake here.
> Would I do it now, share my proposals, No.
> Would I do it if funders’ rules required it. Yes.
> When might funders’ rules require it eg when Tax payers insist that the
> priority is achieving societal goals asap. Might that happen in the
> foreseeable future? I don’t think so because we are as scientists good at
> thinking so far out of the box, such as the internet, or from the 19the
> century electricity and magnetism, the tax payer sees the benefit of an
> individual’s curiosity driven research.
> The bigger point is can we also think beyond individual nations?
> We know we can: the UN, International Council for Science, IUCr……
> So, it probably isn’t a one size fits all idea that James has put forward…
> Best wishes,
> John
>
>
>
> Emeritus Professor John R Helliwell DSc
>
>
>
>
> On 27 Jun 2022, at 19:03, Debanu Das  wrote:
>
> 
> >So, 2nd question is: would you do it? Would you upload your application
> >into the public domain for all to see? What about the reviewer comments?
> >If not, why not?  Afraid people will steal your ideas? Well, once
> >something is public, its pretty clear who got the idea first.
>
> I do not think this ("upload your application into the public domain for
> all to see") is a workable or desirable idea for a variety of reasons.
> There are far greater issues that just about getting credit for your ideas.
> Which is somewhat of an academic and personal pursuit.
>
> For one, the entire R paradigm and programs and IP of entire nations
> (which seems primarily would be the US and potentially some EU countries
> under this case who if at all choose to sign up for this), universities,
> companies (business grants) and funding agencies will wreak havoc (~30-40%
> of US GDP). We already know there is a lopsided distribution of which
> countries taxpayers are funding major IP & innovation. So there are major
> economic, political, social and national competitiveness aspects at stake.
> I doubt that even NSF, DoD, DOE, NIH/HHS or any other government funding
> agency will support such initiatives. Transparency and openness in
> publishing research is a different ball game, even though there too there
> are lopsided effects at the end in many cases, but overall good for world
> progress, hopefully.
>
> Best,
> Debanu
>
> On Wed, Jun 22, 2022 at 6:09 PM James Holton  wrote:
>
>> Greetings all,
>>
>> I'd like to ask a question that I expect might generate some spirited
>> discussion.
>>
>> We have seen recently a groundswell of support for openness and
>> transparency in peer review. Not only are pre-prints popular, but we are
>> also seeing reviewer comments getting published along with the papers
>> themselves. Sometimes even signed by the reviewers, who would have
>> traditionally remained anonymous.
>>
>> My question is: why don't we also do this for grant proposals?
>>
>> I know this is not the norm. However, after thinking about it, why
>> wouldn't we want 

Re: [ccp4bb] open review?

[Debanu Das]

>So, 2nd question is: would you do it? Would you upload your application
>into the public domain for all to see? What about the reviewer comments?
>If not, why not?  Afraid people will steal your ideas? Well, once
>something is public, its pretty clear who got the idea first.

I do not think this ("upload your application into the public domain for
all to see") is a workable or desirable idea for a variety of reasons.
There are far greater issues that just about getting credit for your ideas.
Which is somewhat of an academic and personal pursuit.

For one, the entire R paradigm and programs and IP of entire nations
(which seems primarily would be the US and potentially some EU countries
under this case who if at all choose to sign up for this), universities,
companies (business grants) and funding agencies will wreak havoc (~30-40%
of US GDP). We already know there is a lopsided distribution of which
countries taxpayers are funding major IP & innovation. So there are major
economic, political, social and national competitiveness aspects at stake.
I doubt that even NSF, DoD, DOE, NIH/HHS or any other government funding
agency will support such initiatives. Transparency and openness in
publishing research is a different ball game, even though there too there
are lopsided effects at the end in many cases, but overall good for world
progress, hopefully.

Best,
Debanu

On Wed, Jun 22, 2022 at 6:09 PM James Holton  wrote:

> Greetings all,
>
> I'd like to ask a question that I expect might generate some spirited
> discussion.
>
> We have seen recently a groundswell of support for openness and
> transparency in peer review. Not only are pre-prints popular, but we are
> also seeing reviewer comments getting published along with the papers
> themselves. Sometimes even signed by the reviewers, who would have
> traditionally remained anonymous.
>
> My question is: why don't we also do this for grant proposals?
>
> I know this is not the norm. However, after thinking about it, why
> wouldn't we want the process of how funding is awarded in science to be
> at least as transparent as the process of publishing the results? Not
> that the current process isn't transparent, but it could be more so.
> What if applications, and their reviewer comments, were made public?
> Perhaps after an embargo period?  There could be great benefits here.
> New investigators especially, would have a much clearer picture of
> format, audience, context and convention. I expect unsuccessful
> applications might be even more valuable than successful ones. And yet,
> in reality, those old proposals and especially the comments almost never
> see the light of day. Monumental amounts of work goes into them, on both
> sides, but then get tucked away into the darkest corners of our hard
> drives.
>
> So, 2nd question is: would you do it? Would you upload your application
> into the public domain for all to see? What about the reviewer comments?
> If not, why not?  Afraid people will steal your ideas? Well, once
> something is public, its pretty clear who got the idea first.
>
> 3rd question: what if the service were semi-private? and you got to get
> comments on your proposal before submitting it to your funding agency?
> Would that be helpful? What if in exchange for that service you had to
> review 2-3 other applications?  Would that be worth it?
>
> Or, perhaps, I'm being far too naiive about all this. For all I know
> there are some rules against doing this I'm not aware of.  Either way,
> I'm interested in what this community thinks. Please share your views!
> On- or off-list is fine.
>
> -James Holton
> MAD Scientist
>
> 
>
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1
>
> This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a
> mailing list hosted by www.jiscmail.ac.uk, terms & conditions are
> available at https://www.jiscmail.ac.uk/policyandsecurity/
>



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1

This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing list 
hosted by www.jiscmail.ac.uk, terms & conditions are available at 
https://www.jiscmail.ac.uk/policyandsecurity/

Re: [ccp4bb] JCSG structure validation server

[Debanu Das]

Hi Mike,
As an ex-JCSG team member, I can verify that the JCSG QC structure
validation server (as well as other ones we had), are unfortunately not
operational anymore. We are trying to rebuild (and reinvent along the way)
some more of the JCSG legacy stuff in our current efforts.
Best,
Debanu

On Wed, Dec 1, 2021 at 6:13 AM Hough, Michael  wrote:

> Hi all,
>
>
>
> I recently tried to use the structure validation server and found that the
> web link is no longer active. I wonder if anyone has an idea of whether it
> is still available elsewhere or whether it has been discontinued?
>
>
>
> Thanks,
>
>
>
> Mike
>
> --
>
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1
>



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1

This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing list 
hosted by www.jiscmail.ac.uk, terms & conditions are available at 
https://www.jiscmail.ac.uk/policyandsecurity/

Re: [ccp4bb] looking for proteins with no homologues in pdb

[Debanu Das]

You can download such proteins/sequences from the CASP website:
https://predictioncenter.org/

At the JCSG, we participated for many years in the annual CASP competition,
where we would use/hold our novel experimental crystal structures (before
PDB deposition, almost all our ~1500 targets/PDB depositions had < 20%
sequence identity to existing PDB entries, from novel Pfam/CATH) as tests
by protein structure prediction/modeling groups around the world
participating in CASP to try to predict before PDB release, thereby
allowing a direct comparison with the prediction algorithm and the
experimental structure.

Best,
Debanu
--
Debanu Das

On Mon, Jun 7, 2021 at 1:11 PM Scott Horowitz  wrote:

> For testing purposes, we want to solve structures of proteins that are not
> in the PDB and have no significant sequence homologues in the PDB (i.e. a
> blast of the pdb will get no significant hits). Does anyone happen know a
> good way to find such proteins efficiently? Having an interesting function
> isn't needed.
>
> Thanks,
> Scott
>
> Scott Horowitz, Ph.D.
>
> Assistant Professor
>
> Department of Chemistry & Biochemistry
>
> Knoebel Institute for Healthy Aging
>
> University of Denver
>
>
>
> ECS Building
>
> 2155 E. Wesley Ave
>
> Denver, CO 80208
>
> Phone: 303-871-4326
>
> Fax: 303-871-7915
>
> Zoom Room: https://udenver.zoom.us/my/scotthorowitz
>
> Email: scott.horow...@du.edu
>
> Office: Room 561   Lab: Room 505
>
>
> --
>
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1
>



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1

This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing list 
hosted by www.jiscmail.ac.uk, terms & conditions are available at 
https://www.jiscmail.ac.uk/policyandsecurity/

Re: [ccp4bb] writing coordinates of full biomol into one (PDB) file

[Debanu Das]

And to run it as a single command line option:
http://legacy.ccp4.ac.uk/html/pisa.html

On Tue, May 25, 2021 at 1:07 PM Debanu Das  wrote:

> Easy way to do it online: https://www.ebi.ac.uk/pdbe/pisa/
> or here: http://www.ccp4.ac.uk/pisa/
>
> Best,
> Debanu
>
> On Tue, May 25, 2021 at 1:04 PM Frank Von Delft <
> frank.vonde...@cmd.ox.ac.uk> wrote:
>
>> Yes I thought so too, but discovered I am too stupid to decode the highly
>> parsimonious manual on the ccp4 pages.
>>
>> What I need is a command line that always works. Presumably that's a well
>> defined problem...?
>>
>>
>>
>> Sent from tiny silly touch screen <http://www.9folders.com/>
>> --
>> *From:* Debanu Das 
>> *Sent:* Tuesday, 25 May 2021 20:55
>> *To:* Frank Von Delft
>> *Cc:* CCP4BB@JISCMAIL.AC.UK
>> *Subject:* Re: [ccp4bb] writing coordinates of full biomol into one
>> (PDB) file
>>
>> Hi Frank,
>> PISA is your friend here.
>> Thanks,
>> Debanu
>>
>> On Tue, May 25, 2021 at 12:44 PM Frank von Delft <
>> frank.vonde...@cmd.ox.ac.uk> wrote:
>>
>>> Hello all - this presumably has a really simple solution:
>>>
>>> For a PDB file with a (correct) biomolecular assembly record (REMARK
>>> 350), what program do I use to generate and write out the coordinates of
>>> the biomolecular assembly (or one of them).
>>>
>>> Thanks
>>> Frank
>>>
>>> 
>>>
>>> To unsubscribe from the CCP4BB list, click the following link:
>>> https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1
>>>
>>> This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a
>>> mailing list hosted by www.jiscmail.ac.uk, terms & conditions are
>>> available at https://www.jiscmail.ac.uk/policyandsecurity/
>>>
>>



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1

This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing list 
hosted by www.jiscmail.ac.uk, terms & conditions are available at 
https://www.jiscmail.ac.uk/policyandsecurity/

Re: [ccp4bb] writing coordinates of full biomol into one (PDB) file

[Debanu Das]

Easy way to do it online: https://www.ebi.ac.uk/pdbe/pisa/
or here: http://www.ccp4.ac.uk/pisa/

Best,
Debanu

On Tue, May 25, 2021 at 1:04 PM Frank Von Delft 
wrote:

> Yes I thought so too, but discovered I am too stupid to decode the highly
> parsimonious manual on the ccp4 pages.
>
> What I need is a command line that always works. Presumably that's a well
> defined problem...?
>
>
>
> Sent from tiny silly touch screen <http://www.9folders.com/>
> ----------
> *From:* Debanu Das 
> *Sent:* Tuesday, 25 May 2021 20:55
> *To:* Frank Von Delft
> *Cc:* CCP4BB@JISCMAIL.AC.UK
> *Subject:* Re: [ccp4bb] writing coordinates of full biomol into one (PDB)
> file
>
> Hi Frank,
> PISA is your friend here.
> Thanks,
> Debanu
>
> On Tue, May 25, 2021 at 12:44 PM Frank von Delft <
> frank.vonde...@cmd.ox.ac.uk> wrote:
>
>> Hello all - this presumably has a really simple solution:
>>
>> For a PDB file with a (correct) biomolecular assembly record (REMARK
>> 350), what program do I use to generate and write out the coordinates of
>> the biomolecular assembly (or one of them).
>>
>> Thanks
>> Frank
>>
>> 
>>
>> To unsubscribe from the CCP4BB list, click the following link:
>> https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1
>>
>> This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a
>> mailing list hosted by www.jiscmail.ac.uk, terms & conditions are
>> available at https://www.jiscmail.ac.uk/policyandsecurity/
>>
>



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1

This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing list 
hosted by www.jiscmail.ac.uk, terms & conditions are available at 
https://www.jiscmail.ac.uk/policyandsecurity/

Re: [ccp4bb] writing coordinates of full biomol into one (PDB) file

[Debanu Das]

Hi Frank,
PISA is your friend here.
Thanks,
Debanu

On Tue, May 25, 2021 at 12:44 PM Frank von Delft <
frank.vonde...@cmd.ox.ac.uk> wrote:

> Hello all - this presumably has a really simple solution:
>
> For a PDB file with a (correct) biomolecular assembly record (REMARK
> 350), what program do I use to generate and write out the coordinates of
> the biomolecular assembly (or one of them).
>
> Thanks
> Frank
>
> 
>
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1
>
> This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a
> mailing list hosted by www.jiscmail.ac.uk, terms & conditions are
> available at https://www.jiscmail.ac.uk/policyandsecurity/
>



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1

This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing list 
hosted by www.jiscmail.ac.uk, terms & conditions are available at 
https://www.jiscmail.ac.uk/policyandsecurity/

Re: [ccp4bb] unknown density

[Debanu Das]

Hi,

This may be of interest in this discussion (the abstract is enlightening in
itself):

"Water polygons in high‐resolution protein crystal structures"
Jonas Lee, Sung‐Hou Kim
https://pubmed.ncbi.nlm.nih.gov/19551896/

Download software here (you can analyze input pdb file to look at water
polygon structures): https://sourceforge.net/projects/pdbwaterpolygon/

Thanks,
Debanu

On Sun, Apr 4, 2021 at 10:27 AM Debanu  wrote:

> Hi,
>
> This may be of interest in this discussion (the abstract is enlightening
> in itself):
>
> "Water polygons in high‐resolution protein crystal structures"
> Jonas Lee, Sung‐Hou Kim
> https://pubmed.ncbi.nlm.nih.gov/19551896/
>
> Download software here (you can analyze input pdb file to look at water
> polygon structures): https://sourceforge.net/projects/pdbwaterpolygon/
>
> Thanks,
> Debanu
>
> On Tue, Mar 23, 2021 at 2:15 AM Barone, Matthias 
> wrote:
>
>> can confirm jon´s comment. I find these in virtually every high-res
>> structure. some of them wobble a bit given the AA close by, such as Arg.
>>
>>
>> Dr. Matthias Barone
>>
>> AG Kuehne, Rational Drug Design
>>
>> Leibniz-Forschungsinstitut für Molekulare Pharmakologie (FMP)
>> Robert-Rössle-Strasse 10
>> 13125 Berlin
>>
>> Germany
>> Phone: +49 (0)30 94793-284
>> --
>> *From:* CCP4 bulletin board  on behalf of Jon
>> Cooper <488a26d62010-dmarc-requ...@jiscmail.ac.uk>
>> *Sent:* Monday, March 22, 2021 5:38:40 PM
>> *To:* CCP4BB@JISCMAIL.AC.UK
>> *Subject:* Re: [ccp4bb] unknown density
>>
>> Definitely water pentamer, no doubt at all ;-0
>> Cheers, Jon.C.
>>
>> Sent from ProtonMail mobile
>>
>>
>>
>>  Original Message 
>> On 22 Mar 2021, 14:16, Mark J. van Raaij < mjvanra...@cnb.csic.es>
>> wrote:
>>
>>
>> The ring looks too big to be imidazole or a nucleotide or a carbohydrate,
>> so it’s probably mainly water molecules.
>> Perhaps partially replaced by PEG to explain the density between them
>> (i.e. water molecules in most copies of the protein and PEG in some other
>> copies). I’ve seen horse-shoe shaped PEG in a high-res structure before,
>> PEGs in several confirmations might explain a circle.
>> Practically speaking, I’d first model five waters and see if they refine
>> well.
>>
>> Mark
>>
>> On 22 Mar 2021, at 14:58, Sam Tang  wrote:
>>
>> Hello fellow colleagues
>>
>> Hope you are all well while the pandemics persists. I just wonder if
>> anyone may have an idea what this density (looking like a pentagon) might
>> be. The data was collected to 1.8 A and crystal was grown in Bis-tris +
>> PEG3350. Imidazole residual? Nucleotide (the protein itself is
>> nucleotide-binding, but shouldn't be at this particular site)?
>>
>>
>> https://drive.google.com/file/d/1L9UBFmW72P214itM2HJR_DVy3FaA6FEZ/view?usp=sharing
>>
>> Thanks!
>>
>> BRS
>>
>> Sam
>>
>> --
>>
>> To unsubscribe from the CCP4BB list, click the following link:
>> https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1
>>
>>
>>



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1

This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing list 
hosted by www.jiscmail.ac.uk, terms & conditions are available at 
https://www.jiscmail.ac.uk/policyandsecurity/

Re: [ccp4bb] Downloading older CCP4 version

[Debanu Das]

Hi All,
No more responses required, thank you.
Best,
Debanu

On Thu, Oct 8, 2020 at 8:54 AM Debanu Das  wrote:

> Hello CCP4BB-ers,
> Posting this here as I have been unable to get a response yet from the
> CCP4 help desk:
> Can anyone point to how (specific download link for this version) I can
> download ccp4 7.0.72?
> It appears challenging now to access legacy CCP4 versions.
> Thanks,
> Debanu
>



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1

This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing list 
hosted by www.jiscmail.ac.uk, terms & conditions are available at 
https://www.jiscmail.ac.uk/policyandsecurity/

[ccp4bb] Downloading older CCP4 version

[Debanu Das]

Hello CCP4BB-ers,
Posting this here as I have been unable to get a response yet from the CCP4
help desk:
Can anyone point to how (specific download link for this version) I can
download ccp4 7.0.72?
It appears challenging now to access legacy CCP4 versions.
Thanks,
Debanu



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1

This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing list 
hosted by www.jiscmail.ac.uk, terms & conditions are available at 
https://www.jiscmail.ac.uk/policyandsecurity/

Re: [ccp4bb] Sad News

[Debanu Das]

It is very saddening to hear this news. It took me a couple of days to wrap
my head around it and write.

Thank you for this obituary Bob, which captures so eloquently the breadth
of all of Ward's remarkable engagements and contributions.

I interacted with him a few times over the course of the Protein Structure
Initiative structural genomics programs at the BSGC and JCSG, and more
recently over the last couple of years as part of current endeavors. His
efforts contributed very significantly to everything I have been doing for
the past 15 years too. It is a tremendous loss but a life very well lived
with many important contributions to remember.

Best regards,
Debanu
--
Debanu Das,
Accelero Biostructures

On Sun, Jul 19, 2020 at 4:55 PM James Holton  wrote:

> Ward was the Program Official for all my grants!  My life will not be
> the same without him.  He was always so supportive and helpful with
> advice on how to navigate the sometimes convoluted system that is the
> NIH.  From the Protein Structure Initiative to today his hard work has
> made possible my entire scientific career.
>
> "missed" doesn't seem to cover it. May your rest be a peaceful one, Ward.
>
> -James Holton
> MAD Scientist
>
> On 7/18/2020 4:36 AM, Sweet, Robert wrote:
> > I'm writing to acknowledge the passing of Ward Smith during the weekend
> of 5 July. Ward got his PhD with Martha Ludwig at U. of Michigan, and then
> came to UCLA in 1977 to join Dave Eisenberg’s group as a postdoc. During
> the course of things, he met Cheryl Janson, a Paul Boyer postdoc, and they
> were married in 1980.  During his time at UCLA Ward became expert in
> operation of the Xuong/Hamlin/Nielsen multi-wire system at UCSD and tutored
> the UCLA users in its use. In 1985 Ward and Cheryl left UCLA and went to
> Monsanto in St. Louis, where Ward worked as a structural biologist. In 1987
> they went to Agouron Pharmaceuticals in San Diego. And then in 1995 went
> cross-country to SmithKline Beecham (which became Glaxo SmithKline, merging
> with GlaxoWellcome).
> >
> > Ward was very involved with getting IMCA set up as a functional facility
> for pharmaceuticals at Argonne as SmithKline's representative. This
> experience gave him significant credibility in synchrotron macromolecular
> crystallography, and in 2003 he joined the GM/CA-CAT beamlines at the APS
> to help Bob Fischetti and others construct that excellent facility. During
> this time Cheryl worked at Shamrock Structures.
> >
> > Ward moved to the NIH headquarters in 2007. There he took some
> responsibility for the Protein Structure Initiative, also playing an
> important role in supporting NIH synchrotron facilities. In 2010 he became
> the branch chief for the Structural Genomics and Proteomics Technology
> Branch in the Division of Cell Biology and Biophysics.  He remained in that
> position through 2017. At the 2018 NIGMS re-organization Ward went to the
> Biophysics, Biomedical Technology, and Computational Biosciences division
> as the branch chief for the Biomedical Technology Branch.
> >
> > Ward helped oversee the big NIH-funded, $45 M construction of three
> major beamlines at NSLS-II, a project called ABBIX that ran 2011-2017. In
> 2017 he became program director for NIH support of structural biology
> beamlines at NSLS-II and other DOE synchrotrons.
> >
> > Many knew Ward for his always calm, reasoned demeanor; he was
> unflappable, resilient, and friendly. He was well read and devoted to his
> family.
> >
> > 
> >Robert M. Sweet   E-Dress:  sw...@bnl.gov
> >Scientific Advisor, CBMS: The Center for BioMolecular
> >  Structure at NSLS-II
> >Photon Sciences and Biology Dept
> >Brookhaven Nat'l Lab.
> >Upton, NY  11973 U.S.A.
> >Phones: 631 344 3401  (Office)
> >  631 338 7302  (Mobile)
> > 
> > 
> >
> > To unsubscribe from the CCP4BB list, click the following link:
> > https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1
> >
> > This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a
> mailing list hosted by www.jiscmail.ac.uk, terms & conditions are
> available at https://www.jiscmail.ac.uk/policyandsecurity/
>
> 
>
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1
>
> This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a
> mailing list

Re: [ccp4bb] Flexible C terminus

[Debanu Das]

Hi Chitra,

To add to the discussion, I can offer an example about obtaining the
structure of a flexible/disordered N-term of a membrane protein.

Previous structures of the full-length multidrug efflux transporter AcrB
(12 TM helices in each protein ~1000 residues, forms a trimer, so 36 TM
helices) were missing the first 6 residues in the N-term in the cytoplasm
but we could determine this structure and look at some interesting
sequence-structure implications of these first 6 residues in our structure:

"Crystal structure of the multidrug efflux transporter AcrB at 3.1 Å
resolution reveals the N-terminal region with conserved amino acids
Debanu Das,* Qian Steven Xu,* Jonas Y. Lee, Irina Ankoudinova, Candice
Huang, Yun Lou, Andy DeGiovanni, Rosalind Kim, and Sung-Hou Kim"

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2023878/

I think there should also be examples of C-term or N-term
expression/purification tags that are ordered in some crystal forms of a
target but disordered in other crystal forms/structures of the same
target/homologs.

Best,
Debanu
--
Debanu Das

On Thu, Mar 12, 2020 at 5:02 AM chitra latka  wrote:

> Dear Klemens,
>
> I am going to setup the crystallisation of the entire protein anyhow. I
> hope I get lucky :)
>
> Thanks
> Chitra
>
> On Thu, Mar 12, 2020 at 5:12 PM Klemens Wild <
> klemens.w...@bzh.uni-heidelberg.de> wrote:
>
>> On 12.03.20 08:53, chitra latka wrote:
>>
>> Dear All,
>>
>> I am working on a protein that has flexible C terminus. None of the
>> available structures even in homologs have density for C term region
>> (around 20 odd residues). All the available pdb entries have missing
>> density for these 20 residues at C terminus.
>>
>> I am going to try my luck crystallising the entire protein in hope of
>> getting density for C term residues as well (Fingers crossed).
>>
>> Has anyone faced a similar problem where they have managed to get density
>> for a flexible terminus successfully?
>>
>> Any suggestions would be appreciated.
>>
>> Cheers !
>>
>> Chitra Latka
>>
>>
>> --
>>
>> To unsubscribe from the CCP4BB list, click the following link:
>> https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1
>>
>> Dear Chitra
>>
>> I would nevertheless try. Sometimes flexible termini fold back either in
>> cis or in trans (crystal packing, a case I just had Yesterday) and you
>> might learn sth important for biological regulation if you are lucky. At
>> the same time I would truncate the terminus and crystallize the globular
>> domain in parallel.
>>
>> Good luck
>>
>> Klemens
>>
>
>
> --
> Regards
> Chitra
>
> --
>
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1
>



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1

Re: [ccp4bb] Searching similar local structural conformations

[Debanu Das]

Hi,
Yes, SPASM works very nicely for doing a comparison like this. See Fig 3 in
this publication of ours where we performed a similar analysis:
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4116655/
Best,
Debanu
--
Debanu Das

On Wed, Jan 8, 2020 at 3:28 PM Rigden, Dan  wrote:

> Dear Lei
>
>
> You can use ASSAM for this
>
>
> http://27.126.156.175/assam/
> https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3394286/
>
> In the past I've also used SPASM locally for this
>
> http://xray.bmc.uu.se/usf/spasm.html
>
> but I think you'll need to generate your own up to date database to search.
>
>
> Best wishes
>
> Dan
>
>
> --
> *From:* CCP4 bulletin board  on behalf of Zheng,
> Lei 
> *Sent:* 08 January 2020 15:57:23
> *To:* CCP4BB@JISCMAIL.AC.UK
> *Subject:* [ccp4bb] Searching similar local structural conformations
>
>
> Dear CCP4ers,
>
>
>
> We identify a potential ion binding site formed by four residues in a
> structure. I want to search if any other structures have a similar local
> residual geometry. Is there any programs to perform such a searching? For
> example, to search in Protein Data Bank using coordinates of the binding
> site residues? I appreciate your suggestions.
>
>
>
> Happy New Year!
>
> Lei
>
>
>
>
>
> --
>
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1
>
> --
>
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1
>



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1

Re: [ccp4bb] Please share your experience about "ugly" crystals showing good diffraction

[Debanu Das]

Hi Anirban,

At JCSG, we subjected ~180,000 crystals from ~3500 unique/novel
proteins/complexes to X-ray diffraction screening, resulting in >1500
novel structures in the PDB at an average resolution ~2.0A. In theory,
if we had the bandwidth now to sort through all that data to pull out
images of the mounted crystals and their diffraction quality, we could
probably get some good analysis done on this.

However, as an alternative, I can certainly provide some examples that
we have dealt with recently that I hope will add substance to the
anecdotes to motivate your trainees!

a) For one protein, our first crystals were oval shaped, without sharp
geometry/edges, much like a pebble. We got a high quality 1.9A
structure out of it. Subsequent optimization led to much nicer looking
crystals with nice shapes and we collected ~50 data sets from them all
resulting in ~2A structures so not much improved compared to the
initial visually poorer crystals.

b) For another protein, crystals grow nicely but some of the crystals
remain suspended in solution whereas other settle to the bottom and
stick. For the ones which stick, some can be dislodged by gentle
prodding with a nylon loop while harvesting and they result in (along
with the ones that are not stuck) in ~1.8-2A structures. For the ones
which are stuck and cannot be gently dislodged, a nudge with a plastic
tip (out of desperation!) is sufficient to dislodge them, and they
retain their nice visual appearance and shape, but have total loss in
diffraction.

c) We see this too for crystals soaked with ligands. In some cases
after soaking the crystals appear fine but suffer in diffraction
quality, and in some cases they appear to have suffered visually but
result in usable data sets/structures.

So at the end of the day there are crystals that appear nice but do
not diffract well and there are crystals that do not appear nice but
lead to good/usable structures. So it's all about inner beauty. Never
give up on crystals/crystal optimization without testing them by
X-rays.

Best,
Debanu

On Thu, Jun 28, 2018 at 5:07 PM, Anirban Banerjee  wrote:
>
> Dear all,
>
> Apologies for the non-CCP4 related question.
>
> If you have concrete experience about visually unappealing, i.e. ugly
> crystals ( your own take on ugly is fine)  diffracting better than
> comparably similar sized nicer looking crystals of the same protein, will
> you please share here ? Even better if that led to a published structure.
> Might you also have pictures ?
>
> We have all heard anecdotes about not using visual appearance to judge the
> quality of crystals as far as their ability to give good diffraction data is
> concerned but I am trying to gather some concrete pointers here to motivate
> trainees.
>
> Thanks very much for any help.
>
> Best,
>
> Anirban
>
> P.S. I know that there is probably a lot of thought and wisdom  on this this
> specific topic but I am really looking for actual experience.
>
> 
>
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1



To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1

Re: [ccp4bb] Off-topic question

[Debanu Das]

Hi,
I was going to suggest the same but since it has already been said, here’s a 
cheeky suggestion: you could try determining the crystal structure of the 
complex and get a direct sequence readout for the nuclei acid.
Best,
Debanu 

> On Mar 5, 2018, at 9:34 PM, Philippe BENAS 
> <0d88e888355a-dmarc-requ...@jiscmail.ac.uk> wrote:
> 
> Hi Jobi,
> 
> Phenol/CHCl3 extraction, iPrOH precipitation and then nucleic acid sequencing.
> 
> Best regards,
> Philippe
>  
> Philippe BENAS, Ph.D.
> 
> Laboratoire de Cristallographie et RMN Biologiques, UMR 8015 CNRS
> Faculté de Pharmacie, Université Paris Descartes
> Case 48
> Av, de l'Observatoire
> F-75270 PARIS cedex 06
> +33.1.5373.1599
> E-mails: philippe.be...@parisdescartes.fr, philippe_be...@yahoo.fr
> URLs: http://lcrbw.pharmacie.univ-paris5.fr/ , 
> http://lcrbw.pharmacie.univ-paris5.fr/spip.php?article18
> 
> 
> 
> De : Jobichen Chacko 
> À : CCP4BB@JISCMAIL.AC.UK 
> Envoyé le : Mardi 6 mars 2018 5h11
> Objet : [ccp4bb] Off-topic question
> 
> Dear All,
> We are trying to identify/sequence a DNA/RNA fragment (around 100bp) which 
> was co-purified along with our protein.
> The expression was done in E.coli.
> Any suggestions on how to do this.
> Thank you.
> Jobi
> 
> 

Re: [ccp4bb] does 12 A diffraction worth optimization

[Debanu Das]

Hi Natalia,

One rule of thumb in crystallography: anything is possible!

We have seen many cases of individual proteins (or their complexes with small 
molecules, proteins or nucleic acids) where we ended up with structures 
starting from low resolution initial crystals. So before taking a decision, I 
would suggest considering the following:

a) How many of these crystals did you screen? If only a very few, you should 
try more.
b) How big are the crystals?
c) Are these diffraction results from home source or synchrotron?
d) Long long were crystals allowed to grow before you harvested?
e) Did you try a few different cryos to verify they don't show any difference 
despite no visible deterioration?
f) How long does it take to harvest and place in cryo? How long are you soaking 
in cryo before freezing?
g) Are there issues in crystals being stuck in the well or cover slide and you 
have to apply force to dislodge?
h) Are you doing single step cryo or multi-step in increasing amounts of cryo?
i) Did you try growing crystals at different temp (4C and 20C?) or growing with 
cryo?
j) Do you know what the construct of this apo protein can diffract to?

If you have tried all the above, you can then consider trying different DNA 
lengths and blunt-end vs overhangs or different protein constructs.

Happy to discuss more about the specific case.

Best,
Debanu 
Accelero Biostructures 


> On Fri, Mar 2, 2018 at 5:34 PM, Natalia O  wrote:
> Hello,
> 
> 
> 
> I got crystals of protein-nucleic acid complex, rod-shape, reproducible,
> don’t visibly get damaged upon freezing; however they gave diffraction only
> to about 12 A. I tried several crystals. My question is whether such
> crystals worth optimization. Clearly a 4A diffracting crystal could
> potentially be optimized to 3 – 2.5A, but if the diffraction that I am
> getting now is 12A it could suggest that the system is so flexible that
> getting to 3A with this crystal form is not possible at all. I just wonder
> if there is any statistics or a rule of thumb about what initial diffraction
> worth optimization?
> 
> 
> Thank you!
> 
> -Natalia
> 
> 

Re: [ccp4bb] Hydrophobic hotspots

[Debanu Das]

Hi,
You can try the following using protein structures:
1) https://sbgrid.org/software/titles/quilt
2) Swiss PDBViewer, detect hydrophobic patches under Tool Surface
3) EBI PISA and look for negative delta G
4) look at protein-protein interaction and interface dbases 
Best,
Debanu 

> On Jul 26, 2017, at 10:05 AM, Hugh Morgan 
> <1103e90ebb53-dmarc-requ...@jiscmail.ac.uk> wrote:
> 
> Hi all, can anyone recommend a program for identifying hydrophobic 
> (aggregation) hotspots using either the amino acid sequence and/or structural 
> data. 
> Thanks in advance for your help
> Hugh
> 
> Ps. Have tried aggrescan but would like to try a few others and compare, 
> ideally a more structural based program.

Re: [ccp4bb] Protein rapidly precipitates when off ice

[Debanu Das]

Hi,

I was in Sung-Hou Kim's group when this work below was performed and
published and I also tried it out on many occasions. Elegant piece of
work and certainly worth trying.

Aside from the suggestions of trying different pH and related optimum
solubility screening and if higher salt and glycerol are not helping
when off ice, you can consider the following:

a) Try not concentrating the protein and/or reducing the expression
levels. Maybe you do not need to have so much protein if it leads to
relatively rapid precipitation.

b) Set up some crystallization screens with the protein before
concentration, especially if the protein is clean enough after Ni-NTA.
We crystallized many proteins with Ni-NTA followed by tag cleavage,
and second IMAC

c) Do a high speed spin of the precipitated sample to remove the
precipitate, and run on a gel to verify sample, estimate concentration
and set up crystallization screens on that. This is related to (a) to
remove excess protein.

d) set up crystallization screens at 4C immediately or over a few
hours if stabilized by higher salt/glycerol and maybe the chemicals in
the crystallization reagents can stabilize the protein.

e) If it is a nucleic acid binding protein, try complexes with nucleic
acid added during purification or right after at 4C. Or try protein
partners or other ligands.

f) is the protein Cys rich? Can you anticipate/estimate or model
surface exposed Cys or S-S bonds? Do you have adequate reducing agent
in the sample?

g) Lastly, more esoteric stuff like construct and vector optimization,
mutations, etc.

I am sure there may be a few other things you can try and there may be
more suggestions here.

Best,
Debanu
--
Debanu Das

On Thu, Jul 13, 2017 at 10:46 PM, Briggs, David C
<david.bri...@imperial.ac.uk> wrote:
> Hi Chris,
>
> What is the theoretical pI of your protein? If it is around pH 7.5, you
> might try gel filtering your protein into a different buffer/pH combination.
> Try changing by at least 1 pH unit in either direction.
>
> If the pI isn't a problem, then you might try try solubility screening as
> outlined...
>
> http://scripts.iucr.org/cgi-bin/paper?dz5020
>
> HTH,
>
> Dave
>
> --
> Dr David C Briggs
> Hohenester Lab
> Department of Life Sciences
> Imperial College London
> UK
> http://about.me/david_briggs
>
> 
> From: CCP4 bulletin board <CCP4BB@JISCMAIL.AC.UK> on behalf of Chris Fage
> <fage...@gmail.com>
> Sent: Thursday, July 13, 2017 11:40:34 PM
> To: CCP4BB@JISCMAIL.AC.UK
> Subject: [ccp4bb] Protein rapidly precipitates when off ice
>
> Dear CCP4BB Community,
>
> This week, I purified a nicely overexpressing protein by Ni-NTA followed by
> gel filtration. In a 4 C centrifuge, I concentrated my gel filtration
> fractions to ~1 mL, transferred the spin filter to ice, and then collected 2
> uL for measurement on the Nanodrop. Sadly, the protein precipitated heavily
> in the pipet tip before I could dispense it onto the Nanodrop pedestal,
> directly adjacent to my ice box. This effect seems to be abated at 4 C, as
> the protein remained stable in cold room-chilled pipet tips. However, the
> protein also precipitated heavily when overnight at 4 C in 1 mL gel
> filtration buffer (150 mM NaCl, 10 mM HEPES pH 7.5), but not overnight at 4
> C in 10 mL Ni-NTA buffer (500 mM NaCl, 30 mM HEPES pH 7.5, 10% glycerol)
> prior to gel filtration. Has anyone experienced and resolved a similar issue
> before? Do any useful additives come to mind?
>
> Things I have tried with the gel filtration sample:
> -Exchanging buffer to restore the salt concentration to Ni-NTA levels (e.g.
> 500 mM).
> -Exchanging buffer to add 10% glycerol.
> -Simply diluting the protein in gel filtration buffer to rule out
> concentration dependence.
>
> In each case, the protein precipitates to a milky solution within about a
> minute of removal from ice (I am working with 20-50 uL volumes in PCR
> tubes).
>
> Many thanks for any suggestions!
>
> Best,
> Chris

Re: [ccp4bb] crystallization optimization

[Debanu Das]

Yes, we have done this (addition of water to dilute screen reagents in the 
well) and also try it now in some cases and in fact, this is also the rationale 
behind Hampton's Crystal Screen Lite.
Best,
Debanu
--
Debanu Das

> On Jul 12, 2017, at 9:01 AM, Alun R Coker <alun.co...@ucl.ac.uk> wrote:
> 
> So, if we have a commercial 96 well screen where more than around 60% of the 
> drops precipitate. It may be worth diluting the whole screen say (30ul screen 
> and 20ul water in each well) and repeating . rather than diluting the 
> protein.
> 
> Has anyone ever tried this?
> 
> All the best,
> 
> Alun
> 
>> On 12/07/17 16:54, Frank von Delft wrote:
>> Yes, exactly.  Thanks for doing the Right Thing and posting the actual 
>> diagram.  
>> 
>>> On 12/07/2017 16:26, Patrick Shaw Stewart wrote:
>>> 
>>> Alun
>>> 
>>> I agree Frank's point is very interesting - and he intriguingly refers us 
>>> to the phase diagram.
>>> 
>>> Is the point that Line A is longer than Line B ?
>>> 
>>> Best wishes
>>> 
>>> Patrick
>>> 
>>> 
>>> 
>>> 
>>> 
>>> 
>>> 
>>> 
>>> 
>>> 
>>> 
>>> On 12 July 2017 at 14:40, Alun R Coker <alun.co...@ucl.ac.uk> wrote:
>>> Hi Everyone,
>>> 
>>> Franks point is really interesting. We routinely reduce the protein 
>>> concentration when we see too many precipitated wells, but we never dilute 
>>> the   screen. Has anyone tried this?
>>> 
>>> All the best,
>>> 
>>> Alun
>>> 
>>>> On 12/07/17 08:48, Frank von Delft wrote:
>>>> The point I was failing to make:  reducing either 
>>>> protein or precipitant concentration will indeed reduce nucleation, but 
>>>> often won't get you bigger or more single crystals:  it will just make the 
>>>> appearance of crystals less reliable.
>>>> The way to get big single reliable crystals is to increase protein and 
>>>> greatly reduce precipitant.  
>>>> (Even better:  do seeding.  Like Vicky said.  Incredible how often people 
>>>> don't bother to do seeding, yet it solves so many problems.)
>>>> 
>>>> phx
>>>> 
>>>>> On 12/07/2017 07:50, Vicky Tsirkone wrote:
>>>>> Dear Frank,
>>>>> 
>>>>> I may see in the attached pic several nucleation points and a 
>>>>> considerable amount of microcrystals. Based to my knowledge decreasing 
>>>>> the concentration of the precipitant and/or the protein concentration 
>>>>> would be a reasonable approach to refine the initial hits. 
>>>>> By checking the diagram as you correctly mentioned you may see that the 
>>>>> fine tuning of protein and precipitant concetration may lead to the 
>>>>> desirable result without reaching the precipitation zone.
>>>>> 
>>>>> Patrick just check your screens. Just a rule of thumb, if you see 
>>>>> precipitation in the ~60% of your drops then you should   
>>>>>   definitely reduce the protein concentration.
>>>>> 
>>>>> ps dont forget to try the streak seeding, as well.
>>>>> 
>>>>> Have a nice day and again good luck.
>>>>> 
>>>>> Vicky
>>>>> 
>>>>> On Wed, Jul 12, 2017 at 8:50 AM, Frank von Delft 
>>>>> <frank.vonde...@sgc.ox.ac.uk> wrote:
>>>>> Actually, you should try increasing the protein concentration - a lot.  
>>>>> But be prepared to drop the precipitant concentration to almost nothing 
>>>>> (1 or 2% isn't "low").  
>>>>> To understand why, look at the phase diagram and what we assume about 
>>>>> vapour diffusion.  (Which I'm assuming is what you're doing.)
>>>>> 
>>>>>> On 12/07/2017 06:28, Vicky Tsirkone wrote:
>>>>>> Dear Patrick,
>>>>>> 
>>>>>> You may reduce the protein concentation, as well.
>>>>>> Another option could be the streak seeding by exploiting the drop of 
>>>>>> your initial condition.
>>>>>> 
>>>>>> Good luck,
>>>>>> 
>>>>>> V.T. 
>>>>>> 
>>>>>> On Mon, Jul 10, 2017 at 7:17 PM, Patrick Shaw Stewar

Re: [ccp4bb] protein precipitation reg

[Debanu Das]

Hi Akila,

In addition to what others have asked about the dialysis buffer, a few
more comments that might help to decide next steps because the
precipitation (note precipitation and aggregation are related but not
synonymous) may be due to several different or related reasons:

1) At what stage are you dialyzing? Is it after SEC? Could your
protein be too concentrated at that point since your yield is high
leading to some precipitation? How severe is the loss?
2) Did you try more purification before dialysis?
3) Are you removing the detergent in the buffer you are dialyzing against?
4) Can you try buffer exchange during concentration instead of dialysis?
5) Try increasing your NaCl concentration and adding 5-10% glycerol to
improve protein solubility.
6) Did you try cleaving the C-term His-tag before dialysis? Did you
try N-term His tag?
7) Do you really need to dialyze? Did you try assays or
crystallization trials with the purification buffer? You can run a SEC
column without imidazole to remove that before crystallization.
8) PSI/SBKB TargetTrack can be great resource to look at
expression/purification protocols for similar proteins:
http://sbkb.org/tt/
9) Also look up the Tb Structural Genomics resource to see if there is
anything on this target or related targets: http://www.webtb.org/

Best,
Debanu

On Wed, Mar 29, 2017 at 6:38 AM, Akilandeswari Gopalan
 wrote:
> Dear all,
> I am a PhD student doing structural studies on a few proteins from
> Mycobacterium tuberculosis. The gene encoding the proteins I work on are
> cloned into pet22b with c terminal His tag. the proteins are expressing
> well. upon purification I am getting good yield of protein but during
> dialysis, the proteins precipitate. Kindly suggest some solutions to avoid
> aggregation. pI of one protein is 9.7 and that of the other is 5.6
> I am using 25mM Tris pH 7.5 and 100 mM NaCl buffer with 5mM
> beta-mercaptoethanol and 0.5% triton x 100 for lysis, the same buffer with
> 20-30mM imidazole for washing and 300mM imidazole for eluting the proteins.
>
> Thank you
> Regards
> Akila
>
> --
> Akilandeswari G
>

Re: [ccp4bb] comparison of different protein states

[Debanu Das]

Dear Guillermo,

I think the referee has a point because you are comparing two
different proteins to propose a model of 2 states even though the
enzymes are highly similar with conservation of key features. With 40%
id/60% similarity, even if core domain structure and length are
conserved, there are likely differences in loops, etc.? Are any such
structural variations in the vicinity of the functional site or in
regions that could impact approach or function of the active site? If
so, the apo and bound forms could also be due to intrinsic differences
between the two orthologs?

In the absence of additional supporting evidence, I suppose one way to
frame it would be to emphasize that you are proposing a model based on
available experimental evidence on closely related proteins and also
admit the limitations and unconfirmed/speculative nature of the
proposed model?

However, I think there may be something more interesting you can do
for supporting your theory in case there are other experimental
structures of closely related members of the same enzyme family. If
so, you could compare in detail additional apo forms of multiple
related enzymes and then point to the complex form as an outlier due
to its structural differences, which then may represent your proposed
model.

In addition, do you have assay/mutational analysis of your protein
compared to the other one? If so, and if they have similar activity
and full conservation of active sites, then that can also be used to
support your theory that the 2 enzymes are structurally and
functionally very similar and so the two structures are representative
of your proposed pathway/model.

I had a similar situation a few years ago in comparative structural
analysis of 2 nucleases. Active site residues were identical with the
same metal and overall architecture was similar despite other
structure/dimer differences. Referee insisted we mutate our active
site residues and compare, which we did.

Hope this helps.
Best,
Debanu
--
Debanu Das,
Accelero Biostructures

On Tue, Jan 31, 2017 at 9:40 PM, Guillermo Montoya
<guillermo.mont...@cpr.ku.dk> wrote:
>
> Dear all,
>
> first of all sorry for this off-topic question. I am requesting your help to
> find some papers  to convince one referee about the comparison
> of two different protein states.
>
> In our manuscript we show the crystal structure of an
> enzyme. This structure represents the enzyme after catalysis in complex with
> the product.
>  In the discussion we have superimposed the enzyme in the apo conformation
> and the enzyme after catalysis in complex with the product
> and we have  commented the conformational changes observed
> between these 2 states to propose a model.
>
> The point of the referee is that this comparison is not valid because the
> enzymes that we used in the comparison belong to different species.
> They are not the same protein.
>
> However, and this is stated by us  in the figs and the manuscript, these two
> proteins are
> 40% identical and 60% conserved, the polypeptide length is the same, and
> the key amino acids and the domain structure are fully conserved. They are
> obviously orthologs.
>
> I´d really appreciate if you can send me some literature/information
> to support our approach
>
> Thanks a lot for your input
>
>
> best
>
>
>
>
> Guillermo Montoya, Prof., Dr.
> Research Director, Protein Structure and Function Programme
> Novo Nordisk Foundation Center for Protein Research
> Faculty of Health and Medical Sciences, University of Copenhagen,
> Blegdamsvej 3A, DK-2200 Copenhagen, Denmark
> web: www.cpr.ku.dk
>
>
>
>

Re: [ccp4bb] Abraham Szöke

[Debanu Das]

Dear Bill,

So sorry to hear this news and the passing of another wonderful
scientist and contributor in our field. It was very interesting
reading your obituary. May his soul rest in peace.

I first heard about John and Abraham as a graduate student and later
on when I joined Sung-Hou for postdoc training. I was interested in
their holographic method for x-ray structure determination as I was
dealing with a case where I had density for the partial molecule from
molecular replacement and experimental structure factors for the
entire protein/DNA complex and was looking at ways to recover density
for the missing portion in addition to experimental phasing.

"Holographic methods in X-ray crystallography. IV. A fast algorithm
and its application to macromolecular crystallography"
http://scripts.iucr.org/cgi-bin/paper?S0108767395002315

Best,
Debanu
---
LinkedIn: www.linkedin.com/in/debanudas
Cal Alumni: cal.berkeley.edu/debanudas


On Sun, Jan 29, 2017 at 8:17 PM, William G. Scott  wrote:
> Dear Colleagues:
>
> Is is with great sadness that I report the recent passing of Abraham Szöke, a 
> friend, colleague, informal mentor, and an exceptionally intelligent and 
> energetic source of boundless intellectual enthusiasm.
>
> Abraham was a physicist at Livermore who helped to design nuclear weapons for 
> a living, but he had many other interests. One of these was X-ray 
> crystallography, and with his wife and collaborator Hanna Szöke, as well as 
> John Somoza, he developed an approach to crystallographic real-space 
> electron-density refinement and optimization, implemented in a program called 
> EDEN, that produces electron density maps with minimal model bias in a robust 
> manner. The source code, along with a brief explanation and extensive user's 
> manual, is freely available:
>
> https://code.google.com/archive/p/edencrystallography/
>
> https://github.com/wgscott/edencrystallography
>
> EDEN is written in C and is easy to compile on any unix platform. A fink 
> package for OS X is also available, as are packages for various linux 
> distributions.
>
> John and I first met Abraham and Hanna Szöke when we were graduate students 
> at Berkeley. A few years later, I had the opportunity to collaborate with 
> them and to put EDEN to the test. A single-particle version of EDEN was also 
> under development, for use with electron microscopy. Abraham had a wide 
> variety of interests in addition to electron density reconstruction, 
> including many novel ideas about the origin of life. He enjoyed a multitude 
> of fruitful and productive collaborations throughout a period most normal 
> people would consider retirement. He passed away on Thursday at age 87 from 
> an opportunistic infection while combatting lymphoma.
>
>
> William G. Scott
> Professor, Department of Chemistry and Biochemistry
> and The Center for the Molecular Biology of RNA
> University of California at Santa Cruz
> Santa Cruz, California 95064
> USA
>
> http://scottlab.ucsc.edu

Re: [ccp4bb] Right Handedness of Density

[Debanu Das]

Hi,
  If you have density for the protein and the DNA and the density for 
the protein is correct and you see left-handed DNA density, then I 
suppose you are seeing Z-DNA?


Doesn't seem like a problem of incorrect enantiomorph if your protein 
density is fine. You can pull out a Z-DNA structure from the PDB and see 
if you can get a fit into the density that you see and if it makes sense 
you can then try to build an idealized Z-DNA based on the sequence you 
used for crystallization.


Regards,
Debanu.

Green, Todd wrote:


Hello Ruchi,

I know that when you use non-crystallographic averaging there is a 
possibility of the starting with positive density and ending up with 
negative, the enantiomorph, or the negative enantiomorph density.  In 
order to shift back to the correct phases you can apply a simple 
formula. This is the math that i recall:


positive  structure  alpha
negative structure   alpha + 180
enantiomorphic structure-alpha
negative enantiomorphic structure   -alpha + 180

The recollection comes from a paper in methods in enzymology that i 
don't recall the author or year of at the moment.


I know your case has nothing to do with averaging but the math should 
hold true. In this case, you can switch between the enantiomorphs by 
multiplying the phases by -1. I don't know if this holds a solution to 
your case. It may be worth a quick shot just to multiply the phase 
column by -1. I may be far off base here but i think i am correct. 
This of course is assuming that you have the correct space group and 
that you have the enantiomorphic phases.


If i am totally off on this, someone please correct me. I'm really 
sticking my neck out late on a friday, eh!


Good Luck-
todd


-Original Message-
From: CCP4 bulletin board on behalf of Ruchi Anand
Sent: Fri 3/30/2007 5:31 PM
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] Right Handedness of Density

Dear All,
We are working with a DNA binding protein with 4 Zn sites in in ASU. 
Using SnB and then CNS we were able to get a map using MAD phasing and 
could visualize the density for the double helix of the DNA but it was 
a left handed spiral instead of the usual right handed one. The space 
group which yielded the result was P3(2). We tried to flip the sites 
and change the space group to P3(1) but we are not able to generate a 
sensible map. Any advice regarding the best way to proceed will be 
very helpful. The resolution is around 3.2.

Thanks
Ruchi


This email was scanned with Mcafee's Anti-Virus appliance, but this
is no guarantee that no virus exists. You are asked to make sure you
have virus protection and that it is up to date.

Re: [ccp4bb] Decent dataset from tiny needle crystals--but can't refine past a certain point

[Debanu Das]

Hi Peter,
   After you have finished checking the data again for twinning, etc. 
as suggested by Eleanor,

you might try the following:
1) Refine using the simulated annealing refinement protocols in CNS and 
then check R-values and map quality. If you try that, you can also test 
a few different starting temperature values and slow cooling/constant 
cooling during the annealing procedure. Following this, you can generate 
composite omit maps and also try Primer and Switch phasing (look up 
RESOLVE manual) to try to reduce model bias and try to improve map 
quality which may allow you to trace more of the model.


2) What is the sequence identity of the target compared to the 
homologous search model? Have you already changed all the sequence of 
the MR model solution to reflect your actual target sequence? This may 
help to improve your R-values.


3) Seems that you may have several different homologous models to choose 
from. You may re-try MR using different homologs and see which gives you 
the best starting R/Rfree and select that and change as many amino acids 
as you can to reflect your actual target sequence and then try 
refinement again.


4) Use some homology modeling program and try to remove elements of 
secondary structure from your MR solution that may potentially not be in 
your actual target or may be unstructured (insertions, deletions, etc.)
  
It is not extremely unusual to get a MR solution using a dimeric search 
model instead of the individual monomers. You may also  have the R 
values  given the 40% completeness and also the fact that the search 
model is of a homolog.
Hope some of this may help your case although it may still take cycles 
of iterative manual building.


Else, since you have 60% of the model missing, you may just have to go 
for experimental phasing. Since you have a starting MR solution and thus 
initial partial phases, even if you can get one or more low/medium 
resolution derivatives on your needle crystals, you can use difference 
Fourier methods to supplement the partial MR phases with experimental 
phases for a complete structure determination. Of couse, if you are able 
to improve your crystals, you should have more robust crystals for heavy 
atoms soaks.


Thanks,
Debanu.

Eleanor Dodson wrote:


I guess I would go back to the data.

Could there be twinning?
Is the a pseudo translation vector which means there are 
systematically weak zones of data? ( hklview might show something 
strange)


Rfree seems rather close to R  to me for this resolution - maybe it 
should go up a bit?
How many cycles have you done already?  PHaser refines without taking 
FreeR into account I think..

And is there density for the missing bits?

Eleanor

Peter J Stogios wrote:


Hi,

Long-time reader, first-time writer to ccp4bb.

I'm having a problem with a 3.0 angstrom dataset (2.8 if I push it).  
These crystals were thin but very long needles and I could see 
diffraction only at the synchrotron.  Diffraction spots were very 
very small but well defined.  High level of radiation damage.  
P2(1)2(1)2(1), no large unit cell axes.  R-int 7%(14 in highest 
shell), I/sigma 14(8), completeness 98%.  Nothing weird so far and 
the stats actually look good.  I was ecstatic these tiny needles 
diffracted at all!


I expect 2 chains by Matthew's coefficient and 50% solvent.  
Biological unit is a dimer, homologs are dimers, this protein 
purifies as a dimer.  I'm solving it by molecular replacement, using 
Phaser,  which will give a refine-able solution only when searching 
with dimeric models from homologous proteins.  I cannot get solutions 
that make sense or give Z-scores higher than 5 when searching for two 
chains.  But that doesn't matter, I get a solution when I search for 
a dimer that is able to refine.


So far so good.  Here's where I get stuck: refinement with Refmac 
goes well until R/Rfree values of 32/35, but I cannot break this 
barrier.  In fact, building into positive Fo-Fc peaks results in 
R-free getting worse--actually any further refinement at all results 
in R-free going up.  The model is only 40% complete and has many 
missing regions, not only in loops in turns.  I just can't improve 
the model anymore.  I've tried rebuilding in resolve, Arp/warp, and 
of course lots of manual building.


I don't think my data is as bad as to restrict my refinement, so I'm 
confused as to why I've hit this barrier.  Hopefully this description 
isn't too vague but I appreciate any help in advance and I can 
elaborate if you're willing to help!!



~
Peter J Stogios
Ph.D. candidate, Privé Lab
Dept. of Medical Biophysics, University of Toronto
Toronto Medical Discoveries Tower (TMDT) at MaRS
101 College St., Rm. 4-308
Toronto, Ontario M5G 1L7

e: [EMAIL PROTECTED]
w: http://xtal.uhnres.utoronto.ca/prive
p: (416) 581-8550 ext. 7543

Re: [ccp4bb] Multi-copies in one assymetric unit

[Debanu Das]

Hi,
  Do you have any biochemical evidence that points to oligomeric nature 
of your protein? Does SEC indicate monomer/dimer/trimer/tetramer?
If not or even if so, what does the crystal packing with your MR 
solution with 4 molecules suggest about monomer interactions? Does that 
correlate with SEC results?


Assuming your SG is correct, were you doing your MR search with a 
monomer? Does the packing with 4 molecules leave lots of space that can 
be potentially occupied by more monomers?


/If so, and there is indication of dimer or trimer interactions, you 
could try your MR again and let's say search for 3 dimers  or 2 trimers  
or 2 tetramers/.


Also, what program are you using for MR and what is the resolution range 
for the Rot/Trans search?
If you are not using PHASER, you could /try some different variations of 
resolution range for the search and see if you pick up more molecules/.


What kind of refinement did you do after finding the 4 molecules? Have 
you tried simulated annealing refinement and then inspected the 
resulting maps at low sigma level to see evidence for additional molecules?
Also, how much sequence identity do you have between your target and 
search model? That can affect your map quality and R-factors quite a bit.


Regards,
Debanu.

Yanming Zhang wrote:


Sorry I forget to tell ya the details:

The SG of my Data is cubic P4332. Cell is 251.34A in all three 
dimensions.

Resolution is 3.1A. 5,6,7,8 might be the possible copies from Matthew
coeficient. But I just trust 6 because Chinese like the number 6
(happy Chinese new year by the way :)). No pseudo translation judged 
by Native Patterson. Self-rotation at the section kappa=180 shows 
strong peaks at phi=45 and psi=45.At the stage of map generation, the 
Rfree is 42% R is 38%.

Thanks!
Yanming


On Tue, 13 Feb 2007, Eleanor Dodson wrote:

Sometimes this sort of disorder is due to an error , so the first 
thing is to check very carefully that the solution makes sense.


Why are you so sure there are 6 copies in the asymmetric unit?

In situations like this I first worry about SG.

Is there a pseudo-translation vector? This can make it hard to decide 
on the SG..


Is there an alternate spacegroup with fewer molecules in the asymm unit?

What does the self rotation show?



Yanming Zhang wrote:


Dear All,

Maybe this is a trivial question:

My data should have 6 molecules in one assymetric unit.  MR could 
find out 4 molecules. After this, no matter how hard I have tried, 
no more molecules can be found. At this stage, I suppose that all 
other copies are dis-ordered. And go ahead to do refinement with 4 
molecules (ABCD) available.
The density for A is quite good. But for BCD are very dis-ordered. 
Many breaks in chains. I'd like to ask you:

In a situation like this, should I:
A, Use NCS for all copies?
B Do not use NCS at all?
C, Use NCS just for BCD? (even dis-ordered, but similar)?
What is the trick to lower down Rfree as soon as possible, if you 
have experienced the same situation before?



Thanks

Yanming