- phenixbb - phenix-online.org

Refinement differences?
by Mark A Saper 11 Feb '16

11 Feb '16

Hi, I used Phenix, build 2037 to refine a a new structure. I reran the refinement (with the identical parameters) in build 2155 (v1.10.1) and obtained different statistics and slightly worse geometry. Any idea why? In 2037: start final --------------------------------------- R-work: 0.2488 0.2244 R-free: 0.3003 0.2851 RMS(angles): 1.67 0.80 RMS(bonds): 0.020 0.004 Ramachandran outliers: 1.2% (Goal: < 0.2%) Ramachandran favored: 94.5% (Goal: > 98%) Rotamer outliers: 8.7% (Goal: 1%) C-beta outliers: 0 (Goal: 0) Clashscore: 3.14 Overall score: 2.19 2155 start final --------------------------------------- R-work: 0.2488 0.2331 R-free: 0.3003 0.2856 RMS(angles): 1.77 0.72 RMS(bonds): 0.020 0.004 Ramachandran outliers: 1.4% (Goal: < 0.2%) Ramachandran favored: 93.4% (Goal: > 98%) Rotamer outliers: 9.4% (Goal: 1%) C-beta outliers: 0 (Goal: 0) Clashscore: 4.10 Overall score: 2.36 _________________________________ Mark A. Saper, Ph.D. Associate Professor of Biological Chemistry, University of Michigan Ann Arbor MI 48109-1055 U.S.A. saper(a)umich.edu phone (734) 764-3353 fax (734) 764-3323

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differentiate Glu/Gln and Asp/Asn from Electron Density?
by Kaushik Hatti 10 Feb '16

10 Feb '16

Hello, In continuation to my earlier thread (subject: sequence independent model building possible?), I have built model into density without knowledge of sequence for a data diffracted to 1.9A resolution. Current R/Rfree is 15/19, phaser error=16.88 degrees with no Ramachandran outliers. Is there a way we can differentiate between Glu/Gln, Asp/Asn and sometimes Thr/Val directly from density? I have considered the local environment (hydrophobic/hydrophilic/polarity pockets, possible hydrogen bonds/other interactions, buried/exposed, etc...) in choosing one over the other confusing pairs of amino acids. However, I am not absolutely certain in many places. A BLAST of this sequence against all non-redundant protein sequence database yield highest hit of 80% sequence identity. Hence, we are still not sure of sequence of the contaminant protein which got crystallised and want to decipher sequence directly from the structure. Thanks for any pointers/suggestions, Regards, Kaushik -- Stupidity is everyone’s birthright. However, only the learned exercise it! --Kaushik (28Oct2014)

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Creating cyclic peptide coordinates and restraints
by A User 10 Feb '16

10 Feb '16

Hi Everyone, I am working on a 1.9A enzyme inhibitor complex. The inhibitor is a macrocyclic peptide with a non-proteinogenic amino acid that is modified at the n-terminus with a chloroacetyl-group that cyclizes via the sulfhydryl group of the next cysteine. The total lenght of the peptide is 9 amino acids. What is the most straight forward way to generate the coordinates of such macrocyclic peptide and what are the best settings in eLBOW to generate the restraints for it? Thanks for your time guys! Ben

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"Use starting structure as reference" for homo-multimers
by Tristan Croll 10 Feb '16

10 Feb '16

Hi, I've noticed that when I choose the option to use the starting structure as a reference model and my structure is a homo-multimeric protein, Phenix uses only the first chain as the reference for all chains during refinement. Personally I'd much prefer it if instead it ran a true 1:1 reference throughout the structure, since I tend to use this for final very fine-scale refinement of structures where I've already identified and modelled differences between NCS copies. Others' mileage may vary, of course. Best regards, Tristan

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Re: [phenixbb] Refinement differences?
by Mark A Saper 10 Feb '16

10 Feb '16

Hi Eckhard, Thanks for commenting on this. Unfortunately the data is not good, so I am cutting it off at 2.6 Å resolution. I probably won’t completely refine this structure. It does have 2 molecules in different conformations in the asymmetric unit so I could try NCS. -Mark On Feb 10, 2016, at 10:51 AM, phenixbb-request(a)phenix-online.org wrote: > From: Eckhard Hofmann <eckhard.hofmann(a)bph.ruhr-uni-bochum.de> > Subject: Re: [phenixbb] Refinement differences? > Date: February 10, 2016 at 10:51:08 AM EST > To: phenixbb(a)phenix-online.org > > > Dear Mark, > there were chances to the geometry restraints which most likely would affect your refinement. > Do you have NCS in your refinement? This has been overhauled completely, and is currently not very verbose. > Generally I would not be very concerned about the exact values, given changes in the program. And it looks to me that you are not yet at the final point of refinement anyway. > What is your resolution? > All the best, > Eckhard

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Computational Crystallography Newsletter, Volume 7, Number 1
by Nigel Moriarty 09 Feb '16

09 Feb '16

I am pleased to announce the publication of the latest issue of the Computational Crystallography Newsletter: http://www.phenix-online.org/newsletter/ A listing of the articles and short communications is given below. Please note that the newsletter accepts articles of a general nature of interest to all crystallographers. Please send any articles to me at NWMoriarty(a)lbl.gov noting that there is a Word Template on the website to streamline production. January 2016 <http://www.phenix-online.org/newsletter/CCN_2016_01.pdf> Short Communications - Characterization of base pair geometry <http://www.phenix-online.org/newsletter/CCN_2016_01.pdf#page=5> - Visual representations of internal coordinate restraints: Advantages and limitations <http://www.phenix-online.org/newsletter/CCN_2016_01.pdf#page=9> Fitting tips #11 – Can a helical DNA basepair be Hoogsteen? <http://www.phenix-online.org/newsletter/CCN_2016_01.pdf#page=3> Cheers Nigel --- Nigel W. Moriarty Building 33R0349, 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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Trouble with occupancy refinement
by Chris Fage 08 Feb '16

08 Feb '16

Hello, I've built several glucose molecules from my cryosolution into regions of weaker electron density. Attempting to refine the occupancies of the corresponding atoms, I set the value of each to 0.25 in the pdb file. However, after running phenix.refine, most of the atoms still have occupancies of 1.00: 14 atoms with occ = 0.00 5 atoms with 0.00 < occ < 1.00 74 atoms with occ = 1.00 Is this unusual? Thank you! Best, Chris

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Re: [phenixbb] sequence independent model building possible?
by Kay Diederichs 08 Feb '16

08 Feb '16

To check if you crystallized a contaminant, use https://strube.cbrc.kaust.edu.sa/contaminer/ HTH, Kay > >On 02/05/2016 01:01 AM, Kaushik Hatti wrote: >> Hello, >> >> Is abinitio model building possible for a map with poly alanine model >at 1.9A resolution? >> >> We thought we had crystallised our protein of interest X, collected >data at 1.9 A and all attempts to solve protein X (which has many >homologs) through MR failed. All attempts to re-crystallise the same >protein also failed. >> >> Now, we think the initial protein which got crystallised could be a >contaminant (we don't have any crystals left from this batch to check >for the sequence of the crystallised protein). Through various methods >(and a bit of luck) we have arrived at a decent map with LLG : 3600 and >TFZ: 22 and R/Rfree : 37/41 (for a poly alanine model). >> >> I believe these scores indicate right fold. As I still don't know >the sequence information, is it possible to build sidechains directly >from the map (I could only identify a couple of residues and the model >largely remains PolyAla)? Autobuild with Rebuild-in-place didn't help >in identifying any more residues. >> >> I have also searched PDB database for similar structures. But, none >of those are either from our expression system (E. coli) or organism of >our protein of interest. Neither did I find any similar sequences from >E. coli or our organism of interest. >> >> Any leads/suggestions would be helpful. >> Thanks, >> Kaushik, >> MRN Murthy lab, >> MBU, >> IISc, India >> >> -- > -- Diese Nachricht wurde von meinem Android-Mobiltelefon mit K-9 Mail gesendet.

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Dual conformation on symmetry axis
by Jan Abendroth 06 Feb '16

06 Feb '16

Hi all, I am running into the following problem during the refinement of a rather small structure at 2.15Å resolution: An Arg side chain is located very close to a crystallographic 2-fold. There is good evidence that the side chain has two conformations. However, because of the 2-fold, conformer A overlaps with its symmetry mate A*, as does conformer B. Phenix wants to avoid this 'clash' and moves A and A* out of the density, same for B and B*. Is there a way to avoid this without reducing the symmetry? In the real crystal, of course the two side chains are agnostic as to if they are called A or B and will avoid one another ... Thanks! Jan -- Jan Abendroth Emerald Biostructures Seattle / Bainbridge Island, WA, USA home: Jan.Abendroth_at_gmail.com work: JAbendroth_at_embios.com http://www.emeraldbiostructures.com

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sequence independent model building possible?
by Kaushik Hatti 05 Feb '16

05 Feb '16

Hello, Is abinitio model building possible for a map with poly alanine model at 1.9A resolution? We thought we had crystallised our protein of interest X, collected data at 1.9 A and all attempts to solve protein X (which has many homologs) through MR failed. All attempts to re-crystallise the same protein also failed. Now, we think the initial protein which got crystallised could be a contaminant (we don't have any crystals left from this batch to check for the sequence of the crystallised protein). Through various methods (and a bit of luck) we have arrived at a decent map with LLG : 3600 and TFZ: 22 and R/Rfree : 37/41 (for a poly alanine model). I believe these scores indicate right fold. As I still don't know the sequence information, is it possible to build sidechains directly from the map (I could only identify a couple of residues and the model largely remains PolyAla)? Autobuild with Rebuild-in-place didn't help in identifying any more residues. I have also searched PDB database for similar structures. But, none of those are either from our expression system (E. coli) or organism of our protein of interest. Neither did I find any similar sequences from E. coli or our organism of interest. Any leads/suggestions would be helpful. Thanks, Kaushik, MRN Murthy lab, MBU, IISc, India -- Stupidity is everyone’s birthright. However, only the learned exercise it! --Kaushik (28Oct2014)

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