- phenixbb - phenix-online.org

Re: [phenixbb] Scaling of diffraction data
by jens j birktoft 02 May '12

02 May '12

Hi Everybody, Is there a protocol for re-scaling several sets of diffraction data against a common reference set, either native or based on a Fcalc set computed from a common input model. The are from a number of derivatives, are all isomorphus, and are anomalous data . We want to compare the relative peak heights in Fo-Fc and Anom. difference maps Cheers Jens +++++++++++++++++++++++++++++++++++++++++++++++++++ Jens J. Birktoft e-mail: birktoft @ nyu.edu <jens.knold(a)gmail.com> ; Phone: 212-749-5057 +++++++++++++++++++++++++++++++++++++++++++++++++++

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Xtriage problem
by Abhinav Kumar 02 May '12

02 May '12

Hi, When I try to do R vs R analysis for calculated data using xtriage, it has a problem accepting calc labels. source /home/sw/rhel5/x86_64/phenix/phenix-1.7.3-928/phenix_env phenix.xtriage_1.7.3-928 junk.mtz calc_labels=FC ==> Sorry: No matching array: scaling.input.xray_data.calc_labels=FC Howevere, it works fine with an earlier version: source /home/sw/rhel5/x86_64/phenix/phenix-1.6.4-486/phenix_env phenix.xtriage_1.6.4-486 junk.mtz calc_labels=FC Thanks, Abhinav JCSG@SSRL, SLAC (650) 926-2992 -------- Original Message -------- Subject: RE: SP16569N-180314 Date: Tue, 1 May 2012 15:09:00 -0700 From: Miller, Mitchell D. <mmiller(a)slac.stanford.edu> To: Kumar, Abhinav <abhinavk(a)slac.stanford.edu> Hmm, it seems like it may be a bug introduced into phenix. (or else they changed they syntax and did not update the documentation) Can you report it? I get it to do the RvR analysis for calculated data if I use source /home/sw/rhel5/x86_64/phenix/phenix-1.6.4-486/phenix_env phenix.xtriage_1.6.4-486 junk.mtz calc_labels=FC but if I use source /home/sw/rhel5/x86_64/phenix/phenix-1.7.1-743/phenix_env phenix.xtriage_1.7.1-743 junk.mtz calc_labels=FC or source /home/sw/rhel5/x86_64/phenix/phenix-1.7.3-928/phenix_env phenix.xtriage_1.7.3-928 junk.mtz calc_labels=FC then I get the error Sorry: No matching array: scaling.input.xray_data.calc_labels=FC Thanks, Mitch P.S. the results I got were: R vs R statistic: R_abs_twin =<|I1-I2|>/<|I1+I2|> Lebedev, Vagin, Murshudov. Acta Cryst. (2006). D62, 83-95 R_abs_twin observed data : 0.451 R_abs_twin calculated data : 0.502 R_sq_twin =<(I1-I2)^2>/<(I1+I2)^2> R_sq_twin observed data : 0.279 R_sq_twin calculated data : 0.335 The analyses of the Patterson function reveals a significant off-origin peak that is 44.82 % of the origin peak, indicating pseudo translational symmetry. The chance of finding a peak of this or larger height by random in a structure without pseudo translational symmetry is equal to the 1.4447e-04. See the relevant section of the logfile for more details. The results of the L-test indicate that the intensity statistics behave as expected. No twinning is suspected. Even though no twinning is suspected, it might be worthwhile carrying out a refinement using a dedicated twin target anyway, as twinned structures with low twin fractions are difficult to distinguish from non-twinned structures. -----Original Message----- From: Kumar, Abhinav Sent: Tuesday, May 01, 2012 1:30 PM To: Miller, Mitchell D. Subject: Re: SP16569N-180314 I am having trouble inputting Calc labels to xtriage: My mtz file has the following columns: FreeR_flag FP SIGFP FC PHIC FC_ALL PHIC_ALL FWT PHWT DELFWT PHDELWT FOM PHCOMB I tried "FC,PHIC" and "FC" for calc labels but xtriage does not like it. It writes: No matching array: scaling.input.xray_data.calc_labels=FC Possible choices: /mnt/exdsstaff/abhinavk/Refine/SP16569N-180314/RESULTS/current.mtz:FP,SIGFP /mnt/exdsstaff/abhinavk/Refine/SP16569N-180314/RESULTS/current.mtz:FOM /mnt/exdsstaff/abhinavk/Refine/SP16569N-180314/RESULTS/current.mtz:PHCOMB Thanks, Abhinav JCSG@SSRL, SLAC (650) 926-2992 On 05/01/2012 12:51 PM, Miller, Mitchell D. wrote: Also, you could try running xtriage with FC's from the untwined refinement to look at the R vs R stats for Fc vs Fobs.

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Re: [phenixbb] Geometry Restraints - Anisotropic truncation
by Frank von Delft 01 May '12

01 May '12

Hi Pavel > we discussed this with Randy the other day.. A couple of copy-pasts from > that discussion: > > In general, given highly anisotropic data set: > > 1) maps calculated using all (unmodified) data by phenix.refine, > phenix.maps and similar tools are better than maps calculated using > anisotropy truncated data. So, yes, for the purpose of map calculation > there is no need to do anything: Phenix map calculation tools deal with > anisotropy very well. How did you define "better"? If there are a lot of reflections without signal, that makes them essentially missing, so by including them, you're effectively filling in for those reflections with only DFc. If anisotropy is very strong (i.e. many missing reflections), does that not introduce very significant model bias? The maps would look cleaner, though. (I haven't tested myself, btw; I'm genuinely wondering.) > 2) phenix.refine refinement may fail if one uses original anisotropic > data set. This is probably because the ML target does not use > experimental sigmas (and anisotropy correction by UCLA server is nothing > but Miller index dependent removing the data by sigma criterion - yeah, > that old well criticized practice of throwing away the data that you > worked hard to measure!). May be using sigmas in ML calculation could > solve the problem but that has to be proved. If there are large swathes of reciprocal space without signal - I don't see why shouldn't that be excluded? Tossing out individual reflections is of course silly, but what's wrong with trimming down the edges of the elipsoid? Cheers phx

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Inputting DNA for Phenix MR and/or Autobuild
by Charles Allerston 01 May '12

01 May '12

Dear all, I can't find much in the way of documentation for this. Can someone point me to some please? I basically want to know the correct format for entering DNA into the search for MR and inputting into Autobuild. If I have a double stranded DNA do I enter it as >DNA1 AATT:AATT >DNA1 AATT TTAA etc. ? cheers, charlie Dr. Charles Allerston Genome Integrity & Repair Structural Genomics Consortium Nuffield Department of Medicine Old Road Campus University of Oxford OX3 7DQ http://www.thesgc.org/scientists/groups/oxford/genome-integrity-and-repair

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Call for submissions to June 2012 Computational Crystallography Newsletter (CCN)
by Nigel Moriarty 01 May '12

01 May '12

Calling for articles and short communications of interest to structural biologists. The deadline for publication in the June 2012 issue is 29 May, 2012. The Computational Crystallography Newsletter (CCN) is an electronic newsletter for structural biologists, and is published online every 6 months. Feature articles, meeting announcements and reports can be submitted to me at any time for consideration. Submission of text by email or word-processing files using the CCN templates is requested. Past newsletters and the template are available at www.phenix-online.org/newsletter. Cheers Nigel -- Nigel W. Moriarty Building 64R0246B, Physical Biosciences Division Lawrence Berkeley National Laboratory Berkeley, CA 94720-8235 Phone : 510-486-5709 Email : NWMoriarty(a)LBL.gov Fax : 510-486-5909 Web : CCI.LBL.gov

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Estimation of ligand binding affinity by occupancy refinement
by Zoran Stefanic 01 May '12

01 May '12

Dear all, I have a question about sensitivity of occupancy refinement in phenix. I have a series of datasets done at our home instrument of relatively modest resolutions from 2.8 to 3 A. These are performed on crystals soaked in progressively higher ligand concentrations. We are trying to see to which of the several active sites the ligand binds first so we went from low to high ligand concentrations. We wanted to estimate the amount of ligand bound by refining the occupancies of the ligand. We expect one active site to bind more strongly than the other two active sites. This was indeed what we observed but when we refine the occupancies of the ligands in weaker binding sites starting at 0.2 occupancies they rise to 0.5-0.6 although some atoms are totally out of density and some are in. B factors are reasonable, i.e. don't go wild. My question would be: how reliable is this estimation of ligand binding by occupancy refinement? Why doesn't the occupancy of ligands stay low if afterwards the density fit of the ligand looks rather unconvincing? Thank you in advance Zoran -- Dr. Zoran Stefanic Division of Physical Chemistry Rudjer Boskovic Institute Bijenicka 54 P.O. Box 180 Zagreb Croatia Phone: 00 385 1 457 1203 Fax: 00 385 1 468 0245

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metal coordination
by Panne Daniel 30 Apr '12

30 Apr '12

Hi all, I had a question concerning metal coordination (zinc in this case). I have used phenix.metal_coordination to generate geometry restraints for refinement. However phenix.refine does not write out the necessary LINK records into the PDB file. This seems to lead to problems later during validation, as the metal coordination is not properly recognized. What would be the best way to generate the LINK records in Phenix? Thanks, Daniel

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Error of Sculptor
by Zhang yu 26 Apr '12

26 Apr '12

Dear Phenixer, When I used Sculptor to generate a new model, there is a error which I can not figure out. I input a PDB file (*.pdb) and an alignment file (*.aln). The following is the log file (contents in ######). There seems something wrong with the *.aln file. ################# Reading PDB file ---------------- File name: **/**/******/******/**.pdb Separating solvent from macromolecules... Reading alignment files ----------------------- 1. **/**/**.aln ##################### The following is the content of the alignment file: **_** MVYSYTEKKRIRKDFGKRPQVLDVPYLLSIQLDSFQKFIEQDPEGQY----GLEAAFRSV **_** ---------MEIKRFGRIREVIPLPPLTEIQVESYRRALQADVPPEKRENVGIQAAFRET * **: :*: :* * .**::*::: :: * : *::****.. What is the problem with the alignment file? Thank you in advance! Fish

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cut-out mtz edition
by Dialing Pretty 26 Apr '12

26 Apr '12

Dear All, I just cut out a fragment of chain and the corresponding electron density from the whole molecule. Will you please give me some suggestion on how to edit the attached file so that those electron density not from the protein fragment can be erased? Or do you have other methods to produce high quality figure which just shows the fragment chain and the fragment eletron density? Cheers, Dialing

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Geometry Restraints - UPDATE
by Mario Sanches 25 Apr '12

25 Apr '12

Last week I asked a question about geometry restraints at low resolution. I would like to thank you all that took the time to answer, specially Pavel for looking at the data. First, here is the original question: __________________________________________ I am refining a low resolution structure (2.9A) using phenix.refine. As I continue the refinement Rwork/Rfree are dropping but my geometry statistics in general are very bad. My clashscore is getting worst and the RMS(angle) is really high. I have tried to play with the "wxc_scale" parameter and also ran phenix selecting the "Optimize X-ray/ADP weight" option. All attempts end with a RMS(angle) of ~3.8 and a Clashscore of ~190. Is there a known procedure to deal with this? I feel that my geometry weighing term is too loose but I don't know exactly how to tighten it other than decreasing the wxc_scale parameter. I have gone and manually corrected bad positions before but given the low resolution nature of the structure the clashes tend to be reestablished during refinement. Thank you in advance for your help, __________________________________________ It turns out that the reason to my nightmares was a highly anisotropic dataset. When I tried to fix geometry related problems in coot, phenix.refine was twisting the geometry back again, probably because it was trying to fit a lot of noise due to the anisotropy. Everything was getting worst, geometry, clashscore, and Ramachandran. Pavel finally corrected it by using this server: http://services.mbi.ucla.edu/anisoscale/ I should have been more diligent regarding that, since I could clearly see that my data was anisotropic from the diffraction images. I just didn't know how important this parameter could be. Anyways, phenix.xtriage will tell you if your data is anisotropic. The lesson is to pay attention to it. In my case, the quality of the electron density improved considerably, the geometry improved massively (manual intervention in coot was absolutely necessary, but now it stays correct after I manually fix the problems), clashscore lowered, etc. As a side note, it worked better without hydrogens in my case than with. It is certainly a parameter to try if you are having problems. Thank you all again for the tips. Mario Sanches -- Mario Sanches Postdoctoral Researcher Samuel Lunenfeld Research Institute Mount Sinai Hospital 600 University Ave Toronto - Ontario Canada M5G 1X5 http://ca.linkedin.com/in/mariosanches

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labelit.index with compatibility file
by "Weiergräber, Oliver H." 24 Apr '12

24 Apr '12

Hello, I have used labelit.check_pdb_symmetry to evaluate a P1 dataset and model for possible higher symmetry. C2 turned out to be a candidate solution. When I used the new asymmetric unit content file (xyz_coset0.pdb) to force indexing in the new SG: labelit.index /path/to/my/images/template_####.img 1 90 compatibility_file=xyz_coset0.pdb, the program crashes with AttributeError: 'NoneType' object has no attribute 'mosaicity_spot_coverage' This happens with the version included in phenix-1.7.3 as well as with the latest standalone labelit package. Any ideas what might be causing this? Maybe the new cell fails to refine? It is absolutely possible that forcing the higher symmetry SG leads to significant positional rmsd - I actually believe that P1 is correct, but it's definitly not far from C2. Best regards ================================================ PD Dr. Oliver H. Weiergräber Institute of Complex Systems ICS-6: Structural Biochemistry Tel.: +49 2461 61-2028 Fax: +49 2461 61-1448 ================================================ ------------------------------------------------------------------------------------------------ ------------------------------------------------------------------------------------------------ Forschungszentrum Juelich GmbH 52425 Juelich Sitz der Gesellschaft: Juelich Eingetragen im Handelsregister des Amtsgerichts Dueren Nr. HR B 3498 Vorsitzender des Aufsichtsrats: MinDir Dr. Karl Eugen Huthmacher Geschaeftsfuehrung: Prof. Dr. Achim Bachem (Vorsitzender), Karsten Beneke (stellv. Vorsitzender), Prof. Dr.-Ing. Harald Bolt, Prof. Dr. Sebastian M. Schmidt ------------------------------------------------------------------------------------------------ ------------------------------------------------------------------------------------------------ Kennen Sie schon unsere app? http://www.fz-juelich.de/app

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twinning question
by Patrick Loll 20 Apr '12

20 Apr '12

Dumb question here: When one runs the various twinning tests (e.g. phenix.xtriage), is one meant to use data processed in the higher symmetry SG, or the lower symmetry SG? A concrete example: I can process a data set in a 622 point group w/ reasonable merging statistics, but an abnormally low solvent content raises my suspicions. So do I process as P6, and submit these data to the twinning test? Or do I submit the data processed as P622? Pat --------------------------------------------------------------------------------------- Patrick J. Loll, Ph. D. Professor of Biochemistry & Molecular Biology Director, Biochemistry Graduate Program Drexel University College of Medicine Room 10-102 New College Building 245 N. 15th St., Mailstop 497 Philadelphia, PA 19102-1192 USA (215) 762-7706 pat.loll(a)drexelmed.edu

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Geometry Restraints
by Mario Sanches 19 Apr '12

19 Apr '12

I am refining a low resolution structure (2.9A) using phenix.refine. As I continue the refinement Rwork/Rfree are dropping but my geometry statistics in general are very bad. My clashscore is getting worst and the RMS(angle) is really high. I have tried to play with the "wxc_scale" parameter and also ran phenix selecting the "Optimize X-ray/ADP weight" option. All attempts end with a RMS(angle) of ~3.8 and a Clashscore of ~190. Is there a known procedure to deal with this? I feel that my geometry weighing term is too loose but I don't know exactly how to tighten it other than decreasing the wxc_scale parameter. I have gone and manually corrected bad positions before but given the low resolution nature of the structure the clashes tend to be reestablished during refinement. Thank you in advance for your help, -- Mario Sanches Postdoctoral Researcher Samuel Lunenfeld Research Institute Mount Sinai Hospital 600 University Ave Toronto - Ontario Canada M5G 1X5 http://ca.linkedin.com/in/mariosanches

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inverting a set of phase angles
by Mooers, Blaine H.M. (HSC) 19 Apr '12

19 Apr '12

Does phenix have a reflection file utility for correctly inverting a set of phase angles? Blaine Mooers Assistant Professor Department of Biochemistry and Molecular Biology University of Oklahoma Health Sciences Center S.L. Young Biomedical Research Center Rm. 466 Letter address: Shipping address: P.O. Box 26901, BRC 466 975 NE 10th Street, BRC 466 Oklahoma City, OK 73190 Oklahoma City, OK 73104-5419 office: (405) 271-8300 lab: (405) 271-8313 fax: (405) 271-3910 e-mail: blaine-mooers(a)ouhsc.edu<mailto:[email protected]> webpage: http://www.oumedicine.com/department-of-biochemistry-and-molecular-biology/…

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torsion NCS and SA
by dirk.reinert＠boehringer-ingelheim.com 19 Apr '12

19 Apr '12

Dear all, Is it safe to use torsion NCS (refinement.ncs.type="torsion") and torsion SA (refinement.main.simulated_annealing_torsion= "true") as a default? Thanks for advice, Dirk --- Dr. Dirk Reinert Boehringer Ingelheim Pharma GmbH & Co. KG Dept. Lead Identific. and Optim. Sup. Ge Tel.: +49 (7351) 54-97892 Fax: +49 (7351) 83-97892 mailto:[email protected] Boehringer Ingelheim Pharma GmbH & Co. KG, Sitz: Ingelheim am Rhein; Registergericht Mainz: HR A 22206; Komplementär Boehringer Ingelheim Deutschland GmbH; Geschäftsführung: Dr. Engelbert Günster (Vorsitzender), Ursula Fuggis-Hahn, Ralf Gorniak, Michael Klein, Dr. Martin Wanning; Vorsitzender des Aufsichtsrates: Prof. Dr. Dr. Andreas Barner; Sitz: Ingelheim am Rhein; Registergericht Mainz: HR B 23260 Diese E-Mail ist vertraulich zu behandeln. Sie kann besonderem rechtlichen Schutz unterliegen. Wenn Sie nicht der richtige Adressat sind, senden Sie bitte diese E-Mail an den Absender zurück, löschen die eingegangene E-Mail und geben den Inhalt der E-Mail nicht weiter. Jegliche unbefugte Bearbeitung, Nutzung, Vervielfältigung oder Verbreitung ist verboten. / This e-mail is confidential and may also be legally privileged. If you are not the intended recipient please reply to sender, delete the e-mail and do not disclose its contents to any person. Any unauthorized review, use, disclosure, copying or distribution is strictly prohibited.

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Can the Update Waters routine ignore specified waters?
by John Pascal 19 Apr '12

19 Apr '12

Hello All, While using Update Waters in the phenix GUI, several waters that have been defined around a metal binding site are removed. I am happy with the placement of these waters - can I specify somewhere that Update Waters should leave them alone? and that their positions, etc. are still refined during the run? Thanks. Best Regards, John -- *John M. Pascal, PhD* Assistant Professor Department of Molecular Biology & Biochemistry Kimmel Cancer Center Thomas Jefferson University 233 South 10th Street, BLSB Room 804 Philadelphia, PA 19107 Email: John.Pascal(a)KimmelCancerCenter.org <mailto:[email protected]> Phone: (215) 503-4596 Fax: (215) 923-2117

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(no subject)
by Tom Burnley 18 Apr '12

18 Apr '12

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Re: [phenixbb] Bulk Solvent
by Nathaniel Echols 17 Apr '12

17 Apr '12

Garib's response below, since the server wouldn't let him post. ---------- Forwarded message ---------- From: Garib N Murshudov <garib(a)MRC-LMB.CAM.AC.UK> To: "Steiner, Roberto" <roberto.steiner(a)kcl.ac.uk> Cc: PHENIX user mailing list <phenixbb(a)phenix-online.org> Date: Tue, 17 Apr 2012 18:11:24 +0100 Subject: Re: [phenixbb] Bulk Solvent Dear all I cannot say much without data. Only thing I can say that there were some problems in some of the versions in 5.5, and there are some improvements in recent versions. 5.7 series may perform better. Before claiming that improvement is due to particular feature I would do a little bit more analysis. regards Garib On 17 Apr 2012, at 18:03, Steiner, Roberto wrote: difference density on the backbone can also arise (more often) due to suboptimal ADP refinement/restraints..or a bug in a particular build.. best wishes roberto PS I cc-ed this email to Garib in case he's got some info specific for 5.5.0102 On 17 Apr 2012, at 17:22, Timothy Springer wrote: As I said, I cant say that, it is a guess based on two facts. 1) Looking at the output of refinement steps, and seeing that the largest decrease in R during refinement comes in the bulk solvent correction step. 2) It is also based on the common occurrence in the Refmac but not Phenix maps of negative difference density on backbone. This is a symptom of errors in bulk solvent correction. Tim On Apr 17, 2012, at 12:06 PM, Steiner, Roberto wrote: Hi Tim, How can you say the difference is really down to bulk solvent correction? there's several other parameters that affect differences in R factors.... Just curious R On 17 Apr 2012, at 16:27, Timothy Springer wrote: no, it just takes me two years to write up sometimes--- So this is a historical question. But there is a ref in that powerpoint to following ref. If this was implemented in Phenix and not Refmac 2 years ago, I should cite it. Acta Crystallogr D Biol Crystallogr. 2005 Jul;61(Pt 7):850-5. Epub 2005 Jun 24. A robust bulk-solvent correction and anisotropic scaling procedure. Afonine PV, Grosse-Kunstleve RW, Adams PD. Source Lawrence Berkeley National Laboratory, One Cyclotron Road, Building 64R0121, Berkeley, CA 94720, USA. pafonine(a)lbl.gov Tim On Apr 17, 2012, at 11:18 AM, Jeff Headd wrote: Hi Tim, If you are using Phenix 1.5.2, that would not have had the new bulk-solvent model that Pavel is talking about. Are you using that old of a version for your tests? Jeff On Tue, Apr 17, 2012 at 8:07 AM, Pavel Afonine <pafonine(a)lbl.gov> wrote: > > Hi Tim, > > this is because recently (a few weeks ago) we introduced a novel bulk-solvent model and a new overall anisotropic scaling procedure. > > Some detailed overview is here: > > http://www.phenix-online.org/presentations/PhenixSantaFe2012_PA.pdf > > This is going to be published soon. > > Pavel > > > On 4/17/12 7:59 AM, Timothy Springer wrote: > > In one of the structures I have refined, I found phenix 1.5_2 gave 1 to 1.5% lower Rfree and Rwork than REFMAC 5.5.0102. > Can I propose in a publication that "We attribute these differences to methods for handling bulk solvent in PHENIX that are less adversely affected by deficiencies in the crystallographic data."? > This is a guess, because there is a large contribution to refinement from bulk solvent, rather than factual information. I did cross-refinement, and found it going from structures refined in REfmac to Phenix, and vice versa. The Phenix maps looked better, too. > Is this a reasonable suggestion, and would there be a relevant reference? > Tim > > > > _______________________________________________ > phenixbb mailing list > phenixbb(a)phenix-online.org > http://phenix-online.org/mailman/listinfo/phenixbb > _______________________________________________ phenixbb mailing list phenixbb(a)phenix-online.org http://phenix-online.org/mailman/listinfo/phenixbb <ATT00001..c> Roberto Steiner, PhD Group Leader Randall Division of Cell and Molecular Biophysics 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 roberto.steiner(a)kcl.ac.uk _______________________________________________ phenixbb mailing list phenixbb(a)phenix-online.org http://phenix-online.org/mailman/listinfo/phenixbb <ATT00001..c> Roberto Steiner, PhD Group Leader Randall Division of Cell and Molecular Biophysics 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 roberto.steiner(a)kcl.ac.uk Dr Garib N Murshudov Group Leader, MRC Laboratory of Molecular Biology Hills Road Cambridge CB2 0QH UK Email: garib(a)mrc-lmb.cam.ac.uk Web http://www.mrc-lmb.cam.ac.uk

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Bulk Solvent
by Timothy Springer 17 Apr '12

17 Apr '12

In one of the structures I have refined, I found phenix 1.5_2 gave 1 to 1.5% lower Rfree and Rwork than REFMAC 5.5.0102. Can I propose in a publication that "We attribute these differences to methods for handling bulk solvent in PHENIX that are less adversely affected by deficiencies in the crystallographic data."? This is a guess, because there is a large contribution to refinement from bulk solvent, rather than factual information. I did cross-refinement, and found it going from structures refined in REfmac to Phenix, and vice versa. The Phenix maps looked better, too. Is this a reasonable suggestion, and would there be a relevant reference? Tim

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Re: [phenixbb] Sugar woes
by Matthew J. Whitley 17 Apr '12

17 Apr '12

Hi Simon, I have been struggling with very similar issues over the last few weeks, but I (finally!) got it to work appropriately using just the procedure you described. In my case, I was trying to fit and refine a pentasaccharide consisting of MAN and BMA. Adapting what I did to your case, I would suggest the following steps: After loading NAG and MAN monomers into Coot and manually placing them into the electron density in approximately correct positions, save the coordinates and then manually delete the appropriate hydrogens and oxygen from the PDB file. You specifically stated that you did NOT remove the hydrogens, but I found it to be absolutely necessary to delete these atoms, even though my feeling is that Phenix is supposed to remove automatically the appropriate atoms based on the type of glycosidic linkage specified (although I could be wrong on that). In fact, I just deleted all hydrogen atoms from the sugars for purposes of refinement, knowing that I could add them back in later if desired. To determine the appropriate atoms for removal from the coordinate file in order to form the desired glycosidic linkage, run the command phenix.where_mon_lib_list_cif and then look in the specified file for the type of glycosidic linkage you want. For Beta1-2, you get this: BETA1-2 . DEL-HO2 pyranose . DEL-O1 pyranose glycosidic_bond_beta1-2 This means that, in your example, > apply_cif_link { > data_link = "BETA1-2" > residue_selection_1 = chain Z and resname NAG and resid 5 > residue_selection_2 = chain Z and resname MAN and resid 4 > } you should delete the O1 atom from Man 4 of chain Z and the HO2 atom from Nag 5 of chain Z. If the PDB file contains an O1 for Man4 or an HO2 for Nag5, the refinement will fail, and I think that's the cause of the error you're receiving. If everything works as desired, the result should be that C1 of MAN will be connected to O2 of NAG, thus the 1-2 linkage. If you then load the appropriate .CIF files into Coot and/or Phenix, you should be able to refine the sugars as desired. Alpha2-6 links are not included in the mon_lib_list.cif file. If you want to refine such a linkage, my (admittedly limited) understanding is that, unfortunately, you'd have to manually create a CIF file containing the appropriate restraints - bond lengths, angles, etc. Hope you find something useful in all this. I was was ready to pull out my hair several times over the past few weeks, but now that I got it to work, it doesn't seem so bad anymore. Best of luck. Matthew --- Matthew J. Whitley, Ph.D. Postdoctoral Research Fellow Angela Gronenborn Lab Department of Structural Biology University of Pittsburgh School of Medicine On Apr 17, 2012, at 11:00 AM, phenixbb-request(a)phenix-online.org wrote: > > > Message: 6 > Date: Tue, 17 Apr 2012 11:18:57 +0100 > From: Simon Kolstoe <s.kolstoe(a)ucl.ac.uk> > To: PHENIX user mailing list <phenixbb(a)phenix-online.org> > Subject: [phenixbb] Sugar woes > Message-ID: <7BD08A7C-5B94-4448-8E8A-CBA4437E00B6(a)ucl.ac.uk> > Content-Type: text/plain; charset="us-ascii" > > Hi phenixbb, > > Every couple of years I come to build a sugar chain into a protein structure and hope the process is less painful than last time, however it seems things still are not smooth yet! > > I'm trying to build a fairly standard sugar chain onto a protein and am encountering many problems. What I am doing is using coot to get the coordinates for each sugar using the "File-> Get monomer" option and then putting the appropriate link in my phenix.refine .def file after renumbering the sugars. For example: > > apply_cif_link { > data_link = "BETA1-2" > residue_selection_1 = chain Z and resname NAG and resid 5 > residue_selection_2 = chain Z and resname MAN and resid 4 > } > apply_cif_link { > data_link = "BETA1-4" > residue_selection_1 = chain Z and resname GAL and resid 6 > residue_selection_2 = chain Z and resname NAG and resid 5 > } > > For some reason I keep getting the following error: > > --------------- > > Chain: "Z" > Number of atoms: 220 > Number of conformers: 1 > Conformer: "" > Sorry: Traceback (most recent call last): > File "/Applications/PHENIX-1.7.3-928/Contents/phenix-1.7.3-928/cctbx_project/mmtbx/monomer_library/pdb_interpretation.py", line 1042, in apply_mod > mod_mon = self.monomer.apply_mod(mod_mod_id) > File "/Applications/PHENIX-1.7.3-928/Contents/phenix-1.7.3-928/cctbx_project/mmtbx/monomer_library/cif_types.py", line 399, in apply_mod > result.delete_atom_in_place(mod_atom.atom_id) > File "/Applications/PHENIX-1.7.3-928/Contents/phenix-1.7.3-928/cctbx_project/mmtbx/monomer_library/cif_types.py", line 316, in delete_atom_in_place > raise RuntimeError("delete_atom_in_place: unknown atom_id: %s" % atom_id) > RuntimeError: delete_atom_in_place: unknown atom_id: HO2 > apply_mod failure: > pdbres="NAG Z 5 " segid="Z " > comp id: NAG-b-D > mod id: DEL-HO2 > > ------------------ > > I haven't deleted the hydrogens that came with the sugars so what I thought it was doing was trying to delete an atom that isn't there, however adding a dummy atom with the name HO2 didn't work - any ideas? > > Also it seems happy with BETA1-4 links but not ALPHA2-6 links? How do I specify this link? > > Thanks, > > Simon

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Sugar woes
by Simon Kolstoe 17 Apr '12

17 Apr '12

Hi phenixbb, Every couple of years I come to build a sugar chain into a protein structure and hope the process is less painful than last time, however it seems things still are not smooth yet! I'm trying to build a fairly standard sugar chain onto a protein and am encountering many problems. What I am doing is using coot to get the coordinates for each sugar using the "File-> Get monomer" option and then putting the appropriate link in my phenix.refine .def file after renumbering the sugars. For example: apply_cif_link { data_link = "BETA1-2" residue_selection_1 = chain Z and resname NAG and resid 5 residue_selection_2 = chain Z and resname MAN and resid 4 } apply_cif_link { data_link = "BETA1-4" residue_selection_1 = chain Z and resname GAL and resid 6 residue_selection_2 = chain Z and resname NAG and resid 5 } For some reason I keep getting the following error: --------------- Chain: "Z" Number of atoms: 220 Number of conformers: 1 Conformer: "" Sorry: Traceback (most recent call last): File "/Applications/PHENIX-1.7.3-928/Contents/phenix-1.7.3-928/cctbx_project/mmtbx/monomer_library/pdb_interpretation.py", line 1042, in apply_mod mod_mon = self.monomer.apply_mod(mod_mod_id) File "/Applications/PHENIX-1.7.3-928/Contents/phenix-1.7.3-928/cctbx_project/mmtbx/monomer_library/cif_types.py", line 399, in apply_mod result.delete_atom_in_place(mod_atom.atom_id) File "/Applications/PHENIX-1.7.3-928/Contents/phenix-1.7.3-928/cctbx_project/mmtbx/monomer_library/cif_types.py", line 316, in delete_atom_in_place raise RuntimeError("delete_atom_in_place: unknown atom_id: %s" % atom_id) RuntimeError: delete_atom_in_place: unknown atom_id: HO2 apply_mod failure: pdbres="NAG Z 5 " segid="Z " comp id: NAG-b-D mod id: DEL-HO2 ------------------ I haven't deleted the hydrogens that came with the sugars so what I thought it was doing was trying to delete an atom that isn't there, however adding a dummy atom with the name HO2 didn't work - any ideas? Also it seems happy with BETA1-4 links but not ALPHA2-6 links? How do I specify this link? Thanks, Simon

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Phaser-MR issues?
by Noinaj, Nicholas (NIH/NIDDK) [F] 15 Apr '12

15 Apr '12

Hi, I just installed the latest installer (1036) and having issues with Phaser-MR. i have a multi-domain structure (6-domains) and trying to search for the domains individually but starting with the most conserved first using the Direct search method. However, my issues are... 1- Phaser-MR appears to still be using the 'automatically determined search order' instead of the order I have selected despite me having this turned OFF 2- Phaser-MR appears to still be using the 'FAST method' despite me having DIRECT method chosen Anyone else seeing this or could i be doing something wrong here? Thanks in advance for any feedback! Cheers, Nick

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alternate conformation clashes with its symmetry mate
by Page, Rebecca 15 Apr '12

15 Apr '12

Dear phenix BB, I am having an alternate conformation clashing problem that I am having difficulty resolving: I have a protein in which the N-terminal residue is in two conformations.When the residue is in the 'A' conformation, the sidechain overlaps with the 'A' conformation sidechain density of its symmetry mate. I have modeled the occupancy at 50%. However, every time I refine, phenix pushes the 'A' conformer out of the density, likely due to clashing with its symmetry mate. Is there anyway to get around this? Thanks in advance, Rebecca

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help with validating a very low res structure
by Yunji Wu 14 Apr '12

14 Apr '12

Hi there-- Any old school experimental phasing experts want to help me out? I have an 8 angstrom structure solved using molecular replacement, refined to Rwork 0.38 and Rfree of 0.47. I also have a heavy atom (not SeMet) derivative data set to 10 angstroms. The idea is to get as much information from these data as possible so as to validate my current MR model. My approach is to calculate a difference map using the isomorphous difference Fouriers OR anomalous difference Fouriers as the amplitudes, with phases derived from the MR model-- then using the difference map to locate putative heavy atom sites. I would use various controls and experiments to (computationally) validate the credibility of these sites (e.g. cross-phasing with only iso/ano differences). Anyone have any suggestions for how to do this using Phenix? I use the GUI (generally the nightly builds), and am a newcomer to Phenix. Thanks!!

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Re: [phenixbb] on the protonation state of HIS
by Ho Leung Ng 13 Apr '12

13 Apr '12

Hi Dialing, What resolution is your structure? We are developing a method to predict hydrogen atom positions from structures of more modest resolution (out to about 1.7 A). We'd be happy to take a look at the histidines in your structure. Ho Ho Leung Ng University of Hawaii at Manoa Assistant Professor, Department of Chemistry hng(a)hawaii.edu

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