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

2017-01-25 Thread Robbie Joosten 
Hi Tristan,



There are PDB entries that have this, but this makes matters a bit more 
complicated in annotation. You have to define LINKs and leaving atoms. 
Consistency would be nice here, but that is lacking in these entries.



Cheers,

Robbie



Sent from my Windows 10 phone



Van: Tristan Croll<mailto:ti...@cam.ac.uk>
Verzonden: woensdag 25 januari 2017 18:30
Aan: CCP4BB@JISCMAIL.AC.UK<mailto:CCP4BB@JISCMAIL.AC.UK>
Onderwerp: Re: [ccp4bb] Unknown electron density blob, pdb convention for 
partially ordered ligands



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

T



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




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

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

2017-01-25 Thread Tristan Croll 
I've often wondered about PEG (and, I guess, other synthetic polymers): 
wouldn't it just be better to define the monomer, and then model a chain of 
however many monomers you need?

T

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

 

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

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

2017-01-25 Thread Edward A. Berry 

Uma's use of quotes around "di" suggests a related question about PDB 
convention. It was my (perhaps not very good) understanding that ligands should be 
identified by what is actually present in the crystal, and not by what can be modeled. 
For example endogenous ubiquinone is likely to be UQ50 (depending on the species) but 
most of that 50-carbon side chain is hanging out in the lipid or detergent and completely 
disordered. Still we should use the ligand identifier for UQ50, even though codes exist 
for UQ with 5 or 10-carbon side chains that are much better accommodated by the density.

If that is the case, one should not use the pdb identifier for diethylene glycol (PEG) 
when PEG4k was the precipitant, unless you believe that the binding site has specifically 
selected diethylene glycol from an extremely broad range of polymer lengths in the added 
material.  Using the identifier for a much longer PEG will result in a large number of 
"missing atoms" listed in the report, but would eliminate the unreasonable 
assumption that PEG fragment models must always end with a terminal oxygen.

Even if that is the rule, I would agree that PEGs would be a good place to 
ignore the rule. Since PEGs have a MW distribution, it is impossible to know 
exactly what is bound and it may be different in different unit cells. If you 
are not going to get it right no matter what you put, you might as well put 
something that fits.
eab

On 01/25/2017 09:51 AM, Uma Gabale wrote:

Dear all,
Thank you very much for your replies. It is a PEG, a "di"ethylene glycol to be 
precise, in most chains.
Best regards,
Uma.
--
Uma Gabale, PhD
Research Associate
Molecular and Cellular Biochemistry
Indiana University Bloomington

Re: [ccp4bb] Unknown electron density blob

2017-01-25 Thread Uma Gabale 
Dear all,Thank you very much for your replies. It is a PEG, a "di"ethylene 
glycol to be precise, in most chains. Best regards,Uma. --Uma Gabale, 
PhDResearch AssociateMolecular and Cellular Biochemistry
Indiana University Bloomington
 
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Re: [ccp4bb] Unknown electron density blob

2017-01-24 Thread Robbie Joosten 
PEG likes to hang around where it is unwelcome: 
http://onlinelibrary.wiley.com/doi/10.1002/pro.2923/full

Cheers,
Robbie

Sent from my Windows 10 phone

Van: Bernhard Rupp<mailto:hofkristall...@gmail.com>
Verzonden: dinsdag 24 januari 2017 23:20
Aan: CCP4BB@JISCMAIL.AC.UK<mailto:CCP4BB@JISCMAIL.AC.UK>
Onderwerp: Re: [ccp4bb] Unknown electron density blob

Some general remarks about PEG modelling and associated caveats:
http://journals.iucr.org/d/issues/2016/12/00/rr5136/index.html
Section 3.6.5.

Best, BR

From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Ana Luísa 
Moreira de Carvalho
Sent: Tuesday, January 24, 2017 10:11 AM
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] Unknown electron density blob

Just to add to this, in our group, we had a funny case where we found PEG 
around K:

https://drive.google.com/open?id=0B2DrnhrLgvwGSldLVl9pSEptakE
https://drive.google.com/open?id=0B2DrnhrLgvwGZlNiYTFoaktPUHc

Ana Luisa


On 24 Jan 2017, at 17:36, Artem Evdokimov 
<artem.evdoki...@gmail.com<mailto:artem.evdoki...@gmail.com>> wrote:

PEG. It wraps around K or R residues just like you are showing.

Artem
www.harkerbio.com<http://www.harkerbio.com/>
"where every blob has candy inside"

On Jan 24, 2017 12:19 PM, "Uma Gabale" 
<0ebb5dcf3eaa-dmarc-requ...@jiscmail.ac.uk<mailto:0ebb5dcf3eaa-dmarc-requ...@jiscmail.ac.uk>>
 wrote:
Dear all,
While refining a structure at 2.5 A resolution, we observed a 
semi-circular/crescent shaped electron density blob as shown in the attached 
picture. We have been unable to identify it so far, and would appreciate any 
help in identification.
The protein was expressed in E. coli BL21(DE3), purified on Ni-NTA followed by 
gel filtration. The purification buffers included Tris and NaCl (no detergent/ 
other ingredients except for imidazole for Ni-NTA). Crystallization condition 
had HEPES and PEG3350; no cryoprotectant was used.
The blob is surrounded by residues Trp, Thr, Gln, Arg, and Phe.
Thanks and regards,
Uma.

--
Uma Gabale, PhD
Research Associate
Molecular and Cellular Biochemistry
Indiana University Bloomington

Research Assistant Professor at UCIBIO@REQUIMTE-FCT-NOVA
***
Biologia Estrutural - Cristalografia de Raios-X (Gab 6.34)
Dep. Quimica, FCT-UNL
2829-516 Caparica
Portugal
Phone: 00351212948300 (ext: Gab: 10940; Lab: 10962; X-ray Lab: 10915)
Fax: 00351212948550
http://docentes.fct.unl.pt/almc
http://sites.fct.unl.pt/xtal
https://www.facebook.com/XtalNOVA/
***
Single Crystal X-ray Structure Determination Service: 
http://www.dq.fct.unl.pt/en/single-crystal-x-ray-structure-determination

Re: [ccp4bb] Unknown electron density blob

2017-01-24 Thread Bernhard Rupp 
Some general remarks about PEG modelling and associated caveats:

http://journals.iucr.org/d/issues/2016/12/00/rr5136/index.html

Section 3.6.5.

 

Best, BR

 

From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Ana
Luísa Moreira de Carvalho
Sent: Tuesday, January 24, 2017 10:11 AM
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] Unknown electron density blob

 

Just to add to this, in our group, we had a funny case where we found PEG
around K:

 

https://drive.google.com/open?id=0B2DrnhrLgvwGSldLVl9pSEptakE

https://drive.google.com/open?id=0B2DrnhrLgvwGZlNiYTFoaktPUHc

 

Ana Luisa

 

 

On 24 Jan 2017, at 17:36, Artem Evdokimov <artem.evdoki...@gmail.com
<mailto:artem.evdoki...@gmail.com> > wrote:

 

PEG. It wraps around K or R residues just like you are showing.

 

Artem

www.harkerbio.com <http://www.harkerbio.com/> 

"where every blob has candy inside"

 

On Jan 24, 2017 12:19 PM, "Uma Gabale"
<0ebb5dcf3eaa-dmarc-requ...@jiscmail.ac.uk
<mailto:0ebb5dcf3eaa-dmarc-requ...@jiscmail.ac.uk> > wrote:

Dear all,

While refining a structure at 2.5 A resolution, we observed a
semi-circular/crescent shaped electron density blob as shown in the attached
picture. We have been unable to identify it so far, and would appreciate any
help in identification.

The protein was expressed in E. coli BL21(DE3), purified on Ni-NTA followed
by gel filtration. The purification buffers included Tris and NaCl (no
detergent/ other ingredients except for imidazole for Ni-NTA).
Crystallization condition had HEPES and PEG3350; no cryoprotectant was used.

The blob is surrounded by residues Trp, Thr, Gln, Arg, and Phe.

Thanks and regards,

Uma.

 

--

Uma Gabale, PhD

Research Associate

Molecular and Cellular Biochemistry
Indiana University Bloomington

 

Research Assistant Professor at UCIBIO@REQUIMTE-FCT-NOVA

***
Biologia Estrutural - Cristalografia de Raios-X (Gab 6.34)
Dep. Quimica, FCT-UNL
2829-516 Caparica
Portugal
Phone: 00351212948300 (ext: Gab: 10940; Lab: 10962; X-ray Lab: 10915)
Fax: 00351212948550
 <http://docentes.fct.unl.pt/almc> http://docentes.fct.unl.pt/almc
http://sites.fct.unl.pt/xtal

https://www.facebook.com/XtalNOVA/
***

Single Crystal X-ray Structure Determination Service:
http://www.dq.fct.unl.pt/en/single-crystal-x-ray-structure-determination

Re: [ccp4bb] Unknown electron density blob

2017-01-24 Thread Ana Luísa Moreira de Carvalho 
Just to add to this, in our group, we had a funny case where we found PEG 
around K:

https://drive.google.com/open?id=0B2DrnhrLgvwGSldLVl9pSEptakE
https://drive.google.com/open?id=0B2DrnhrLgvwGZlNiYTFoaktPUHc

Ana Luisa


> On 24 Jan 2017, at 17:36, Artem Evdokimov  wrote:
> 
> PEG. It wraps around K or R residues just like you are showing.
> 
> Artem
> www.harkerbio.com 
> "where every blob has candy inside"
> 
> On Jan 24, 2017 12:19 PM, "Uma Gabale" 
> <0ebb5dcf3eaa-dmarc-requ...@jiscmail.ac.uk 
> > wrote:
> Dear all,
> While refining a structure at 2.5 A resolution, we observed a 
> semi-circular/crescent shaped electron density blob as shown in the attached 
> picture. We have been unable to identify it so far, and would appreciate any 
> help in identification.
> The protein was expressed in E. coli BL21(DE3), purified on Ni-NTA followed 
> by gel filtration. The purification buffers included Tris and NaCl (no 
> detergent/ other ingredients except for imidazole for Ni-NTA). 
> Crystallization condition had HEPES and PEG3350; no cryoprotectant was used.
> The blob is surrounded by residues Trp, Thr, Gln, Arg, and Phe.
> Thanks and regards,
> Uma.
> 
> --
> Uma Gabale, PhD
> Research Associate
> Molecular and Cellular Biochemistry
> Indiana University Bloomington

Research Assistant Professor at UCIBIO@REQUIMTE-FCT-NOVA
***
Biologia Estrutural - Cristalografia de Raios-X (Gab 6.34)
Dep. Quimica, FCT-UNL
2829-516 Caparica
Portugal
Phone: 00351212948300 (ext: Gab: 10940; Lab: 10962; X-ray Lab: 10915)
Fax: 00351212948550
http://docentes.fct.unl.pt/almc 
http://sites.fct.unl.pt/xtal
https://www.facebook.com/XtalNOVA/
***
Single Crystal X-ray Structure Determination Service: 
http://www.dq.fct.unl.pt/en/single-crystal-x-ray-structure-determination

Re: [ccp4bb] Unknown electron density blob

2017-01-24 Thread Vivoli, Mirella 
Hi there,

I think it is PEG, very common to have PEG in horseshoe shape with arginine 
pointing toward to it.

Cheers,


Mirella


Vivoli Mirella

Associate Research Fellow
Biocatalysis Centre, Henry Wellcome Building
College of Life and Environmental Sciences,
University of Exeter
Stocker Road
Exeter
Ex4 4QD
Tel: + 44 (0)1392 726121
Email: m.viv...@exeter.ac.uk

"I don't want to believe. I want to know". [C. Sagan]

From: CCP4 bulletin board <CCP4BB@JISCMAIL.AC.UK> on behalf of Paul Emsley 
<pems...@mrc-lmb.cam.ac.uk>
Sent: 24 January 2017 17:32:35
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] Unknown electron density blob

On 24/01/2017 17:19, Uma Gabale wrote:

> While refining a structure at 2.5 A resolution, we observed a 
> semi-circular/crescent shaped
> electron density blob as shown in the attached picture. We have been unable 
> to identify it
> so far, and would appreciate any help in identification.

If you have Coot:

Extensions->Modelling->Add Other Solvent Molecules->Add New Residue 
Type->"PG4"->Add
then click "PG4 Tetraethylene Glycol"

Re: [ccp4bb] Unknown electron density blob

2017-01-24 Thread Artem Evdokimov 
PEG. It wraps around K or R residues just like you are showing.

Artem
www.harkerbio.com
"where every blob has candy inside"

On Jan 24, 2017 12:19 PM, "Uma Gabale" <
0ebb5dcf3eaa-dmarc-requ...@jiscmail.ac.uk> wrote:

> Dear all,
> While refining a structure at 2.5 A resolution, we observed a
> semi-circular/crescent shaped electron density blob as shown in the
> attached picture. We have been unable to identify it so far, and would
> appreciate any help in identification.
> The protein was expressed in *E. coli* BL21(DE3), purified on Ni-NTA
> followed by gel filtration. The purification buffers included Tris and NaCl
> (no detergent/ other ingredients except for imidazole for Ni-NTA).
> Crystallization condition had HEPES and PEG3350; no cryoprotectant was used.
> The blob is surrounded by residues Trp, Thr, Gln, Arg, and Phe.
> Thanks and regards,
> Uma.
>
> --
> Uma Gabale, PhD
> Research Associate
> Molecular and Cellular Biochemistry
> Indiana University Bloomington
>

Re: [ccp4bb] Unknown electron density blob

2017-01-24 Thread Parthasarathy Sampathkumar 
Hi Uma,

It is risky to guess based on one-view of of the density from 2-dimensional
images. What is the contour-level of 2mFo-DFc (blue) map displayed here?!!
If 2mFo-DFc density is continuous from Arg, say at 0.8 sigma, then could it
be an alternate conformation of the Arg side-chain. One could build, and
re-refine and see if negative mFo-DFc features shows up.

Hope this helps,
Partha

On Tue, Jan 24, 2017 at 12:19 PM, Uma Gabale <
0ebb5dcf3eaa-dmarc-requ...@jiscmail.ac.uk> wrote:

> Dear all,
> While refining a structure at 2.5 A resolution, we observed a
> semi-circular/crescent shaped electron density blob as shown in the
> attached picture. We have been unable to identify it so far, and would
> appreciate any help in identification.
> The protein was expressed in *E. coli* BL21(DE3), purified on Ni-NTA
> followed by gel filtration. The purification buffers included Tris and NaCl
> (no detergent/ other ingredients except for imidazole for Ni-NTA).
> Crystallization condition had HEPES and PEG3350; no cryoprotectant was used.
> The blob is surrounded by residues Trp, Thr, Gln, Arg, and Phe.
> Thanks and regards,
> Uma.
>
> --
> Uma Gabale, PhD
> Research Associate
> Molecular and Cellular Biochemistry
> Indiana University Bloomington
>

Re: [ccp4bb] Unknown electron density blob

2017-01-24 Thread Paul Emsley 

On 24/01/2017 17:32, Paul Emsley wrote:

On 24/01/2017 17:19, Uma Gabale wrote:


While refining a structure at 2.5 A resolution, we observed a 
semi-circular/crescent shaped
electron density blob as shown in the attached picture. We have been unable to 
identify it
so far, and would appreciate any help in identification.


If you have Coot:

Extensions->Modelling->Add Other Solvent Molecules->Add New Residue 
Type->"PG4"->Add
then click "PG4 Tetraethylene Glycol"


Delete the water first, obviously.

Re: [ccp4bb] Unknown electron density blob

2017-01-24 Thread Paul Emsley 

On 24/01/2017 17:19, Uma Gabale wrote:


While refining a structure at 2.5 A resolution, we observed a 
semi-circular/crescent shaped
electron density blob as shown in the attached picture. We have been unable to 
identify it
so far, and would appreciate any help in identification.


If you have Coot:

Extensions->Modelling->Add Other Solvent Molecules->Add New Residue 
Type->"PG4"->Add
then click "PG4 Tetraethylene Glycol"

Re: [ccp4bb] Unknown electron density blob

2017-01-24 Thread Boaz Shaanan 



Hi,


Could it be a segment of peg3350?


  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  Skype: boaz.shaanan  
Fax:   972-8-647-2992 or 972-8-646-1710
 
 








From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] on behalf of Uma Gabale [0ebb5dcf3eaa-dmarc-requ...@jiscmail.ac.uk]
Sent: Tuesday, January 24, 2017 7:19 PM
To: CCP4BB@JISCMAIL.AC.UK
Subject: [ccp4bb] Unknown electron density blob





Dear all,
While refining a structure at 2.5 A resolution, we observed a semi-circular/crescent shaped electron density blob as shown in the attached picture. We have been unable to identify it so far, and would
 appreciate any help in identification.
The protein was expressed in
E. coli BL21(DE3), purified on Ni-NTA followed by gel filtration. The purification buffers included Tris and NaCl (no detergent/ other ingredients except for imidazole for Ni-NTA). Crystallization condition
 had HEPES and PEG3350; no cryoprotectant was used.
The blob is surrounded by residues Trp,
Thr,
Gln,
Arg, and Phe.
Thanks and regards,
Uma.



--
Uma Gabale, PhD

Research Associate

Molecular and Cellular Biochemistry
Indiana University Bloomington