High B-factors after Phenix restrained refinement

Da Duan

15 Dec 2011 15 Dec '11

10:20 p.m.

Hi all I am rather new to the crystallography so please bear with me, :-). I used Phenix AutoMR to solved a structure to 3.3A and after 1 round of rigidbody refinement with Phenix Refine I proceeded to restrained refinement. The R/Rfree from the refinement decreased nicely as expected but the B average is at ~100 (using Group B factor refinement option). I took the same model and mtz through Refmac and the B average is about ~40. Has anyone experienced this before? I am almost positive it maybe a setting issue in Phenix Refine that i should be looking at to get the B factors to refine correctly. Thanks in advance for your help! Da Duan

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Pavel Afonine

15 Dec 15 Dec

10:34 p.m.

Hi,

...

I used Phenix AutoMR to solved a structure to 3.3A and after 1 round of rigidbody refinement with Phenix Refine I proceeded to restrained refinement. The R/Rfree from the refinement decreased nicely as expected but the B average is at ~100 (using Group B factor refinement option). I took the same model and mtz through Refmac and the B average is about ~40. Has anyone experienced this before? I am almost positive it maybe a setting issue in Phenix Refine that i should be looking at to get the B factors to refine correctly.

do restrained individual ADP (B-factor) refinement. With the type of restraints that phenix.refine is using it is totally ok at 3.3A resolution. Pavel

Pavel Afonine

10:36 p.m.

New subject: High B-factors after Phenix restrained refinement - p.s.

p.s.: also, make sure you are using the latest PHENIX version. Pavel

Nathaniel Echols

16 Dec 16 Dec

5:31 a.m.

On Thu, Dec 15, 2011 at 2:20 PM, Da Duan <[email protected]> wrote:

...

I used Phenix AutoMR to solved a structure to 3.3A and after 1 round of rigidbody refinement with Phenix Refine I proceeded to restrained refinement. The R/Rfree from the refinement decreased nicely as expected but the B average is at ~100 (using Group B factor refinement option). I took the same model and mtz through Refmac and the B average is about ~40. Has anyone experienced this before? I am almost positive it maybe a setting issue in Phenix Refine that i should be looking at to get the B factors to refine correctly.

Dialing Pretty

8:04 a.m.

Hi Nat, I have a protein-peptide complex crystal, and I got the 3-D structure of the protein part in the complex by rigid body refinement with isomophous crystal crystal structure as the search model, and I got the peptide structure by Coot peptide building, then I refine the whole protein-peptide complex. Will you please tell me which Phenix program can be used to build the peptide as I mentioned in the situation above? I want to get a Coot alternative for the peptide building. I am looking forward to getting your reply. Cheers, Dialing ________________________________ From: Nathaniel Echols To: PHENIX user mailing list Sent: Friday, 16 December 2011 3:31 PM Subject: Re: [phenixbb] High B-factors after Phenix restrained refinement On Thu, Dec 15, 2011 at 2:20 PM, Da Duan <[email protected]> wrote:

...

I used Phenix AutoMR to solved a structure to 3.3A and after 1 round of rigidbody refinement with Phenix Refine I proceeded to restrained refinement. The R/Rfree from the refinement decreased nicely as expected but the B average is at ~100 (using Group B factor refinement option). I took the same model and mtz through Refmac and the B average is about ~40. Has anyone experienced this before? I am almost positive it maybe a setting issue in Phenix Refine that i should be looking at to get the B factors to refine correctly.

How are you calculating the average B? Refmac prints "residual" B-factors in the B column of ATOM records - these do not include the contribution from TLS and Ucryst (an overall B-factor for the entire crystal). In Phenix, the ATOM records always have the total isotropic B-factor, and this will always be higher than the equivalent in Refmac. So it's quite likely that both programs are correct, they're just reporting very different things. (And for what it's worth, a mean B-factor of 100 is totally normal at 3.3A resolution.) -Nat _______________________________________________ phenixbb mailing list [email protected] http://phenix-online.org/mailman/listinfo/phenixbb

Tom Terwilliger

3:32 p.m.

New subject: High B-factors after Phenix restrained refinement -- peptide building

Hi Dialing, You should be able to build the peptide in either of 2 ways: 1. Cut out the density for the peptide with phenix.cut_out_density and build into cutout.mtz with phenix.build_one_model, supplying the sequence of the peptide, and not refining the model. This will just build the peptide. Then you can translate the model that is built back using phenix.cut_out_density. 2. Alternatively, you can put your data file and your map coefficients, and complete sequence file including the peptide into phenix.autobuild and it should try to build the peptide in addition to the rest of your model. You can also include your working model to speed it up. All the best, Tom T On Dec 16, 2011, at 1:04 AM, Dialing Pretty wrote:

...

Hi Nat,

I have a protein-peptide complex crystal, and I got the 3-D structure of the protein part in the complex by rigid body refinement with isomophous crystal crystal structure as the search model, and I got the peptide structure by Coot peptide building, then I refine the whole protein-peptide complex.

Will you please tell me which Phenix program can be used to build the peptide as I mentioned in the situation above? I want to get a Coot alternative for the peptide building.

I am looking forward to getting your reply.

Cheers,

Dialing

From: Nathaniel Echols To: PHENIX user mailing list Sent: Friday, 16 December 2011 3:31 PM Subject: Re: [phenixbb] High B-factors after Phenix restrained refinement

On Thu, Dec 15, 2011 at 2:20 PM, Da Duan <[email protected]> wrote:

...
I used Phenix AutoMR to solved a structure to 3.3A and after 1 round of rigidbody refinement with Phenix Refine I proceeded to restrained refinement. The R/Rfree from the refinement decreased nicely as expected but the B average is at ~100 (using Group B factor refinement option). I took the same model and mtz through Refmac and the B average is about ~40. Has anyone experienced this before? I am almost positive it maybe a setting issue in Phenix Refine that i should be looking at to get the B factors to refine correctly.

How are you calculating the average B? Refmac prints "residual" B-factors in the B column of ATOM records - these do not include the contribution from TLS and Ucryst (an overall B-factor for the entire crystal). In Phenix, the ATOM records always have the total isotropic B-factor, and this will always be higher than the equivalent in Refmac. So it's quite likely that both programs are correct, they're just reporting very different things. (And for what it's worth, a mean B-factor of 100 is totally normal at 3.3A resolution.)

-Nat _______________________________________________ phenixbb mailing list [email protected] http://phenix-online.org/mailman/listinfo/phenixbb

_______________________________________________ phenixbb mailing list [email protected] http://phenix-online.org/mailman/listinfo/phenixbb

Thomas C. Terwilliger Mail Stop M888 Los Alamos National Laboratory Los Alamos, NM 87545 Tel: 505-667-0072 email: [email protected] Fax: 505-665-3024 SOLVE web site: http://solve.lanl.gov PHENIX web site: http:www.phenix-online.org ISFI Integrated Center for Structure and Function Innovation web site: http://techcenter.mbi.ucla.edu TB Structural Genomics Consortium web site: http://www.doe-mbi.ucla.edu/TB CBSS Center for Bio-Security Science web site: http://www.lanl.gov/cbss

Da Duan

3:54 p.m.

Hi Nat I was just looking at the average B in the refinement log files from Refmac and Phenix Refine. Thanks for the clarification on how Refmac and Phenix calculate the average B. My next question is when depositing the structure, is it more common to deposit structures with the "residual" B-factors or B-factors generated by Phenix that includes the TLS and Ucryst contribution? I also performed sfcheck and the average B generated by the Wilson plot is ~100 which seems to suggest that the Phenix average B is probably "more" correct? Thanks again Da On Fri, Dec 16, 2011 at 12:31 AM, Nathaniel Echols wrote:

...

On Thu, Dec 15, 2011 at 2:20 PM, Da Duan <[email protected]> wrote:

...
I used Phenix AutoMR to solved a structure to 3.3A and after 1 round of rigidbody refinement with Phenix Refine I proceeded to restrained refinement. The R/Rfree from the refinement decreased nicely as expected but the B average is at ~100 (using Group B factor refinement option). I took the same model and mtz through Refmac and the B average is about ~40. Has anyone experienced this before? I am almost positive it maybe a setting issue in Phenix Refine that i should be looking at to get the B factors to refine correctly.

How are you calculating the average B? Refmac prints "residual" B-factors in the B column of ATOM records - these do not include the contribution from TLS and Ucryst (an overall B-factor for the entire crystal). In Phenix, the ATOM records always have the total isotropic B-factor, and this will always be higher than the equivalent in Refmac. So it's quite likely that both programs are correct, they're just reporting very different things. (And for what it's worth, a mean B-factor of 100 is totally normal at 3.3A resolution.)

-Nat _______________________________________________ phenixbb mailing list [email protected] http://phenix-online.org/mailman/listinfo/phenixbb

Ed Pozharski

4:05 p.m.

On Fri, 2011-12-16 at 10:54 -0500, Da Duan wrote:

...

is it more common to deposit structures with the "residual" B-factors or B-factors generated by Phenix that includes the TLS and Ucryst contribution?

While this policy http://deposit.rcsb.org/adit/REFMAC.html explicitly mentions refmac, I assume that it is a general expectation of the pdb that you deposit full B-factors. -- Oh, suddenly throwing a giraffe into a volcano to make water is crazy? Julian, King of Lemurs

Nathaniel Echols

4:11 p.m.

On Fri, Dec 16, 2011 at 7:54 AM, Da Duan <[email protected]> wrote:

...

I was just looking at the average B in the refinement log files from Refmac and Phenix Refine. Thanks for the clarification on how Refmac and Phenix calculate the average B. My next question is when depositing the structure, is it more common to deposit structures with the "residual" B-factors or B-factors generated by Phenix that includes the TLS and Ucryst contribution? I also performed sfcheck and the average B generated by the Wilson plot is ~100 which seems to suggest that the Phenix average B is probably "more" correct?

I had thought that the PDB automatically converted Refmac PDB files to contain full B-factors, but the link Ed posted suggests otherwise. If you are depositing the output file from Phenix, at least, you do not need to take any additional steps. -Nat

Steiner, Roberto

4:20 p.m.

Hi Nat The PDBe used to do the conversion automatically (likely still does) After submission they would send you something like this:

...

The PDB no longer accepts or releases entries that contain only residual B-factors. For this reason your entry was converted to full B-factors and ANISOU records encoding the TLS parameters using the protocol as given in:

http://deposit.rcsb.org/adit/REFMAC.html

The converted file was then run against EDS refinement and as no change was observed in the calculated R-factors, this was considered for release.

Best R On 16 Dec 2011, at 16:11, Nathaniel Echols wrote:

...

On Fri, Dec 16, 2011 at 7:54 AM, Da Duan <[email protected]> wrote:

...
I was just looking at the average B in the refinement log files from Refmac and Phenix Refine. Thanks for the clarification on how Refmac and Phenix calculate the average B. My next question is when depositing the structure, is it more common to deposit structures with the "residual" B-factors or B-factors generated by Phenix that includes the TLS and Ucryst contribution? I also performed sfcheck and the average B generated by the Wilson plot is ~100 which seems to suggest that the Phenix average B is probably "more" correct?

I had thought that the PDB automatically converted Refmac PDB files to contain full B-factors, but the link Ed posted suggests otherwise. If you are depositing the output file from Phenix, at least, you do not need to take any additional steps.

-Nat _______________________________________________ phenixbb mailing list [email protected] http://phenix-online.org/mailman/listinfo/phenixbb

Roberto Steiner, PhD Randall Division of Cell and Molecular Biophysics Group Leader King's College London Room 3.10A New Hunt's House Guy's Campus SE1 1UL, London, UK Tel 0044-20-78488216 Fax 0044-20-78486435 [email protected]

Steiner, Roberto

4:11 p.m.

Hi Da Even if you deposit a structure refined with Refmac the PDB now expects the total B values being present. Have a look at http://deposit.rcsb.org/adit/REFMAC.html What you call "more" correct does not really make much sense to me if I understand you properly. If you follow the link given above (or use TLSANL directly from the CCP4) and get 'total Bs' from Refmac I am sure they will be more or less the same and the Bs from phenix.refine. R On 16 Dec 2011, at 15:54, Da Duan wrote: Hi Nat I was just looking at the average B in the refinement log files from Refmac and Phenix Refine. Thanks for the clarification on how Refmac and Phenix calculate the average B. My next question is when depositing the structure, is it more common to deposit structures with the "residual" B-factors or B-factors generated by Phenix that includes the TLS and Ucryst contribution? I also performed sfcheck and the average B generated by the Wilson plot is ~100 which seems to suggest that the Phenix average B is probably "more" correct? Thanks again Da On Fri, Dec 16, 2011 at 12:31 AM, Nathaniel Echols mailto:[email protected]> wrote: On Thu, Dec 15, 2011 at 2:20 PM, Da Duan <[email protected]mailto:[email protected]> wrote:

...

I used Phenix AutoMR to solved a structure to 3.3A and after 1 round of rigidbody refinement with Phenix Refine I proceeded to restrained refinement. The R/Rfree from the refinement decreased nicely as expected but the B average is at ~100 (using Group B factor refinement option). I took the same model and mtz through Refmac and the B average is about ~40. Has anyone experienced this before? I am almost positive it maybe a setting issue in Phenix Refine that i should be looking at to get the B factors to refine correctly.

How are you calculating the average B? Refmac prints "residual" B-factors in the B column of ATOM records - these do not include the contribution from TLS and Ucryst (an overall B-factor for the entire crystal). In Phenix, the ATOM records always have the total isotropic B-factor, and this will always be higher than the equivalent in Refmac. So it's quite likely that both programs are correct, they're just reporting very different things. (And for what it's worth, a mean B-factor of 100 is totally normal at 3.3A resolution.) -Nat _______________________________________________ phenixbb mailing list [email protected]mailto:[email protected] http://phenix-online.org/mailman/listinfo/phenixbb Roberto Steiner, PhD Randall Division of Cell and Molecular Biophysics Group Leader King's College London Room 3.10A New Hunt's House Guy's Campus SE1 1UL, London, UK Tel 0044-20-78488216 Fax 0044-20-78486435 [email protected]mailto:[email protected]

Pavel Afonine

4:56 p.m.

I agree, it is a matter of convention. The total ADP is: Utotal=Ucryst+Ugroup+Ulocal. You can either output the total Utotal into ATOM/ANISOU records or keep Ucryst+Ugroup in REMARKs and output Ulocal into ATOMs. Both ways are valid as long as they yield identical Utotal. For more relevant information, summary and some review, see: - pages 23-29 here: http://phenix-online.org/presentations/latest/pavel_refinement_general.pdf - article "On atomic displacement parameters (ADP) and their parameterization in PHENIX" here: http://phenix-online.org/newsletter/ Pavel On 12/16/11 8:11 AM, Steiner, Roberto wrote:

...

Hi Da

Even if you deposit a structure refined with Refmac the PDB now expects the total B values being present. Have a look at http://deposit.rcsb.org/adit/REFMAC.html

What you call "more" correct does not really make much sense to me if I understand you properly. If you follow the link given above (or use TLSANL directly from the CCP4) and get 'total Bs' from Refmac I am sure they will be more or less the same and the Bs from phenix.refine.

R

On 16 Dec 2011, at 15:54, Da Duan wrote:

...
Hi Nat

I was just looking at the average B in the refinement log files from Refmac and Phenix Refine. Thanks for the clarification on how Refmac and Phenix calculate the average B. My next question is when depositing the structure, is it more common to deposit structures with the "residual" B-factors or B-factors generated by Phenix that includes the TLS and Ucryst contribution? I also performed sfcheck and the average B generated by the Wilson plot is ~100 which seems to suggest that the Phenix average B is probably "more" correct?

Thanks again

Da

On Fri, Dec 16, 2011 at 12:31 AM, Nathaniel Echols mailto:[email protected]> wrote:

On Thu, Dec 15, 2011 at 2:20 PM, Da Duan <[email protected] mailto:[email protected]> wrote: > I used Phenix AutoMR to solved a structure to 3.3A and after 1 round of > rigidbody refinement with Phenix Refine I proceeded to restrained > refinement. The R/Rfree from the refinement decreased nicely as expected but > the B average is at ~100 (using Group B factor refinement option). I took > the same model and mtz through Refmac and the B average is about ~40. Has > anyone experienced this before? I am almost positive it maybe a setting > issue in Phenix Refine that i should be looking at to get the B factors to > refine correctly.

How are you calculating the average B? Refmac prints "residual" B-factors in the B column of ATOM records - these do not include the contribution from TLS and Ucryst (an overall B-factor for the entire crystal). In Phenix, the ATOM records always have the total isotropic B-factor, and this will always be higher than the equivalent in Refmac. So it's quite likely that both programs are correct, they're just reporting very different things. (And for what it's worth, a mean B-factor of 100 is totally normal at 3.3A resolution.)

-Nat _______________________________________________ phenixbb mailing list [email protected] mailto:[email protected] http://phenix-online.org/mailman/listinfo/phenixbb

Roberto Steiner, PhD Randall Division of Cell and Molecular Biophysics Group Leader King's College London

Room 3.10A New Hunt's House Guy's Campus SE1 1UL, London, UK Tel 0044-20-78488216 Fax 0044-20-78486435 [email protected] mailto:[email protected]

_______________________________________________ phenixbb mailing list [email protected] http://phenix-online.org/mailman/listinfo/phenixbb

Da Duan

18 Dec 18 Dec

3:49 p.m.

Thanks to everyone who've replied to this thread. I really appreciated the help and the useful information. Merry Xmas and Happy New Year everyone!! Cheers Da On Fri, Dec 16, 2011 at 11:56 AM, Pavel Afonine wrote:

...

I agree, it is a matter of convention. The total ADP is: Utotal=Ucryst+Ugroup+Ulocal. You can either output the total Utotal into ATOM/ANISOU records or keep Ucryst+Ugroup in REMARKs and output Ulocal into ATOMs. Both ways are valid as long as they yield identical Utotal.

For more relevant information, summary and some review, see:

- pages 23-29 here: http://phenix-online.org/presentations/latest/pavel_refinement_general.pdf

- article "On atomic displacement parameters (ADP) and their parameterization in PHENIX" here: http://phenix-online.org/newsletter/

Pavel

On 12/16/11 8:11 AM, Steiner, Roberto wrote:

Hi Da

Even if you deposit a structure refined with Refmac the PDB now expects the total B values being present. Have a look at http://deposit.rcsb.org/adit/REFMAC.html

What you call "more" correct does not really make much sense to me if I understand you properly. If you follow the link given above (or use TLSANL directly from the CCP4) and get 'total Bs' from Refmac I am sure they will be more or less the same and the Bs from phenix.refine.

R

On 16 Dec 2011, at 15:54, Da Duan wrote:

Hi Nat

I was just looking at the average B in the refinement log files from Refmac and Phenix Refine. Thanks for the clarification on how Refmac and Phenix calculate the average B. My next question is when depositing the structure, is it more common to deposit structures with the "residual" B-factors or B-factors generated by Phenix that includes the TLS and Ucryst contribution? I also performed sfcheck and the average B generated by the Wilson plot is ~100 which seems to suggest that the Phenix average B is probably "more" correct?

Thanks again

Da

On Fri, Dec 16, 2011 at 12:31 AM, Nathaniel Echols wrote:

...
On Thu, Dec 15, 2011 at 2:20 PM, Da Duan <[email protected]> wrote:

...
I used Phenix AutoMR to solved a structure to 3.3A and after 1 round of rigidbody refinement with Phenix Refine I proceeded to restrained refinement. The R/Rfree from the refinement decreased nicely as expected but the B average is at ~100 (using Group B factor refinement option). I took the same model and mtz through Refmac and the B average is about ~40. Has anyone experienced this before? I am almost positive it maybe a setting issue in Phenix Refine that i should be looking at to get the B factors to refine correctly.

How are you calculating the average B? Refmac prints "residual" B-factors in the B column of ATOM records - these do not include the contribution from TLS and Ucryst (an overall B-factor for the entire crystal). In Phenix, the ATOM records always have the total isotropic B-factor, and this will always be higher than the equivalent in Refmac. So it's quite likely that both programs are correct, they're just reporting very different things. (And for what it's worth, a mean B-factor of 100 is totally normal at 3.3A resolution.)

-Nat _______________________________________________ phenixbb mailing list [email protected] http://phenix-online.org/mailman/listinfo/phenixbb

Roberto Steiner, PhD Randall Division of Cell and Molecular Biophysics Group Leader King's College London

Room 3.10A New Hunt's House Guy's Campus SE1 1UL, London, UK Tel 0044-20-78488216 Fax 0044-20-78486435 [email protected]

_______________________________________________ phenixbb mailing [email protected]http://phenix-online.org/mailman/listinfo/phenixbb

_______________________________________________ phenixbb mailing list [email protected] http://phenix-online.org/mailman/listinfo/phenixbb

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Da Duan
Dialing Pretty
Ed Pozharski
Nathaniel Echols
Pavel Afonine
Steiner, Roberto
Tom Terwilliger