Re: [ccp4bb] Bad density for chains

[Pooja Kesari]

Dear All,
Thank you all for reply.

We have checked the data for twinning.
Our protein is 360 residues around 40 kDa protein.
We have tried TLS refinement.
chain A and B don't superimpose well with chain C and D. (A and B chains
also share slight difference )
Since we don't have proper density for *some regions*  chain C and D, we
are not sure whether these chain have similar or different conformations.
We tried anisotropy correction and the model refined a bit.


On Wed, Jan 25, 2017 at 10:32 AM, Debanu  wrote:

> Hi Pooja,
>
> Are you positive you have the correct space group and there are no other
> issues like twinning, etc?
>
> If sure, did you define NCS groups in refinement? TLS refinement? Try
> different refinement programs?
>
> How big is the molecule? Was it solved by MR or experimental phasing?
>
> You can try superimposing A/B on C/D and refinement with tight NCS then
> adjust NCS restraints during model adjustments based on local differences
> or also see if phenix autobuild helps.
>
> Best,
> Debanu
> --
> Debanu Das
> Accelero Biostructures
>
>
> On Jan 24, 2017, at 8:42 PM, Pooja Kesari  wrote:
>
> Dear All,
>
> I have a 2.6 A resolution structure having four chains in an asymmetric
> unit.
> The chain A and B have density for almost all residues however we don't
> have proper residue density in chain C and D.What can be tried to build
> chain C and D ?
>
>
>
> Many Thanks
> Pooja
>
>


-- 
Thanks & Regards,
Pooja Kesari
Research Scholar
Department Of Biotechnology
Indian Institute of Technology Roorkee
INDIA

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

[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
Verzonden: woensdag 25 januari 2017 18:30
Aan: 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  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
>>

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[Ryan Jackson]

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-- 
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Assistant Professor
Dept. Chemistry and Biochemistry
Utah State University
Logan UT, 84322
phone: 435-797-1635
fax:435-797-3390

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

[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

[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] Bad density for chains

[Phoebe A. Rice]

Are you sure there really are 4 chains?  50% solvent may be average, but we've 
had crystals with closer to 80%.



From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] on behalf of Pooja Kesari 
[pkesar...@gmail.com]
Sent: Tuesday, January 24, 2017 10:42 PM
To: CCP4BB@JISCMAIL.AC.UK
Subject: [ccp4bb] Bad density for chains

Dear All,

I have a 2.6 A resolution structure having four chains in an asymmetric unit.
The chain A and B have density for almost all residues however we don't have 
proper residue density in chain C and D.What can be tried to build chain C and 
D ?



Many Thanks
Pooja

[ccp4bb] explain "comfortable"

[Bernhard Rupp]

Hi Fellows,

 

JPK has made a valid point by reminding me that my "statement about becoming
"comfortable" with a particular level of conservatism/enthusiasm"

made in the N14 paper seems like a very non-scientific standard.

 

He is right. Let me explain: The difference between enthusiastic modelling
and being a reckless over-modeller is dependent on your objective, that is,
what

hypothesis or claims is your model at that specific location supposed to
support? This is a question that no statistic or metric is going to answer
for you. 

There is indeed room for interpretation - after all it is *electron density
interpretation* what we do, and some are more perceptive than others; such

is the artistic (or artisan) and perhaps enjoyable part in model building.
As long as the postulate is plausible, you do have a certain degree of
reasonable freedom. 

Solvent for example, is fluid and floating, after all. But also solvent
modeling does have some rules, consequences and meaning.

 

It is important not mistake this disclaimer for a postmodern argument
justifying 'Anything goes'. If one's claim is to support a certain
mechanistic detail or 

ligand pose, one better cough up some convincing, properly generated and
adequately contoured difference electron density and a model that is not
delusional in

view of basic stereochemistry.

Sorry for being redundant.

 

Best, BR

--

Bernhard Rupp

  http://www.hofkristallamt.org/

  b...@hofkristallamt.org

+1 925 209 7429

+43 767 571 0536

--

All models are wrong

but some are useful.

--

 

Re: [ccp4bb] Unknown electron density blob

[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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