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disulfide bonds in phenix.refine
by Allister.Crow＠uea.ac.uk 18 Jan '08

18 Jan '08

Hello all, I am currently trying to refine a structure (with phenix.refine) that contains a number of disulfide bonds. Many of these disulfides appear to be partly broken such that the default restraints given to these disulfides by phenix.refine cause the cysteine sulfur atoms to be placed too close to one another. This results in difference density in the fo-fc map. I want to try to model these partially broken states a bit more accurately, either by modelling two alternative conformations (i.e. one cysteine pair in a disulfide and the other pair as free thiols) or by allowing the disulfide bond distance to stretch. Currently refinement drives the S-S bond length close to dictionary values regardless of how far apart I model the cysteines in my input pdb file and whether the density implies a bridge or not. My two questions are: 1) How do I specifically prevent a disulfide bond from being defined between two sulfurs that are close together (I tried manipulating the "disulfide_distance_cutoff" variable but this doesn't seem to stop a disulfide being formed). 2) How can I soften the restraint on the disulfide bond length for specific cysteine residues. Thanks for your time. Dr. Allister Crow

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Continue from an AutoBuild job abruptly killed
by Miguel Ortiz-Lombardía 17 Jan '08

17 Jan '08

Dear all, We had a problem with a machine running AutoBuild, which resulting in the computer going down and the AutoBuild job consequently killed. I've tried to find out if there is a way to continue the job (it had been running for four days and still had a few hours to go I guess: a pretty big unit cell) but I failed. I would appreciate if someone could point to the relevant pages in the manual/tutorials or give me advice about how to do such a thing. If that's possible, of course. We're using the latest phenix (1.3b-rc6) and cci apps. Thank you! Miguel -- http://www.pangea.org/mol/spip.php?rubrique2 ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ Je suis de la mauvaise herbe, Braves gens, braves gens, Je pousse en liberté Dans les jardins mal fréquentés! Georges Brassens

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MSE gap
by Troy E Messick 16 Jan '08

16 Jan '08

Hello, I seem to be having a problem with phenix.refine and the seleno-methionine residue (resname MSE). It seems that I have a gap in the peptide bond between the C atoms and the N atom of the subsequent residue. Is there something I could add to the restraints_edits.params file to get it to click together? What is the distance_ideal and sigma for a peptide bond? Thanks for your help, Troy

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phenix.find_ncs and DANO
by Bryan W. Lepore 16 Jan '08

16 Jan '08

in principle, phenix.find_ncs should be fine with FP=DANO, right? and - do i understand correctly that phenix only correlates the map with the .pdb sites, and does not rotate the map about itself (like in getax, i think)? -bryan p.s: i can run phenix.find_ncs with FP=DANO, but seems that resolve wants it to be FP as usual.

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autosol : input_ncs_file
by Bryan W. Lepore 16 Jan '08

16 Jan '08

using PHENIX VERSION: 1.3b of 20-11-2007. does phenix.autosol use "input_ncs_file"? the documentation implies that phenix.autosol will take `input_ncs_file` (at the 'find_ncs' entry, "input_ncs_file" is mentioned but no where else), yet, i get a 'Sorry, unknown file or keyword:' message with no such command line option listed. -bryan

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strategy of refinement for a structure phased by bromide soaking
by Pascal EGEA 11 Jan '08

11 Jan '08

Dear All, I have a 1.8A resolution data set of a protein collected on a crystal soaked in KBr and I am solving using Phenix. I used AutoSol and gave a rough estimate of f_prime and f_double_prime for the Br scatterer as a starting point. It worked great and Phenix found 31 Bromide ions (only one is fully occupied) and the resulting map is remarkably clear. Automatic building has done a great job and I was able to build most of the 4 monomers present in the AU. However I am confused and have a couple of questions. 1-As Phenix started it generated a fobs_experimental_phases_free_Rflag.mtz file. I have been using this file and no longer my data.hkl file for further refinement. I would like to know why the best_solution after the automatic building run does not contain any of the Bromide ions. It has water molecules but no Bromide ions? What is the reason for that? 2-I have been refining against the mtz file I mentioned but without any of these Bromide ions and although the refinement is converging , I suspect I have been doing something bad and wrong. Should I start my refinement again from the first best pdb after automatic building and include right away the bromide ions with simultaneous f_prime, f_double_prime and occupancy refinement for the anomalous scatterers? As it stands now the Rfree is 22.9 and the Rfac is 19.7. The map looks fine except for the solvent, some of this atoms are obviously bromide and there is extra density in the Fo-Fc around some "solvent" atoms. Thanks a lot in advance for your suggestions. Pascal Egea University of California San Francisco Department of Biochemistry and Biophysics

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iotbx.shelx.from_ins
by Foelker, Andrew 10 Jan '08

10 Jan '08

Hello Perhaps one of you could assist me with this question. In the from_ins.py file, there is a function by the name of from_ins. I am passing a .RES file to this function, but am not passing any of the other optional arguments to it. When line 51 in the file is reached, an exception is thrown. Here is the line of code that blows up: b = record.tempFactor + record_next.tempFactor The exception correctly states that the record_next object has no attribute named 'tempFactor'. Perhaps I am doing something incorrectly, or perhaps there is a bug in the from_ins function itself. If any of you can offer some insight into this problem I would greatly appreciate it. Thank you. ------------------------------------------------------------------------ -------- Andrew J. Foelker Software Engineer Bruker AXS Inc. 5465 E. Cheryl Parkway Madison, WI 53711 Tel: +1 608-276-3812 andrew.foelker(a)bruker-axs.com www.bruker-axs.com <http://www.bruker-axs.com/> ------------------------------------------------------------------------ ----------

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Reading Rfree Flags from Refmac
by James Fraser 09 Jan '08

09 Jan '08

Hello, Someone in our lab has started a refinement run in REFMAC and wants to continue it in Phenix.refine. Is there an easy way to get phenix to recognize the r-free flag from REFMAC (freeR=0)? Thanks, jf -- James Fraser jamesfraser(a)berkeley.edu Alber Lab 356 Stanley Hall, QB3 UC Berkeley Berkeley, CA 94720 http://ucxray.berkeley.edu/~jfraser/

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Failure when calculating a composite omit map
by Dale Tronrud 07 Jan '08

07 Jan '08

Hi all, I'm trying to calculate a composite omit map so that I can get a clear view of a troublesome region in my structure. I have two copies of everything in the asu so I created some dummy atoms to mask the region in the A chain and called those atoms chain Y and another collection to mask the region in the C chain and created a Z chain for them. I put these atoms in a file name Mask.pdb I attempted to calculate the composite omit map starting from my MR solution but that fails because Autobuild doesn't like being given the Fourier coefficients produced by Phaser. To work around this problem I decided to ask it to create the omit map based on the results of my AutoBuild followed the MR. My script is: phenix.autobuild model=AutoBuild_run_12_/overall_best.pdb data=1M50-2.mtz \ input_refinement_labels="FP SIGFP None None None None None None FreeR_flag" \ seq_file=../fmo-ct.pir \ resolution=2.2 dmax=20 refinement_resolution=2.2 \ cif_def_file_list=/usr/users/dale/geometry/chromophores/bcl_tnt.cif \ input_lig_file_list=AutoMR_run_4_/MR.1-Bchl-a.pdb \ composite_omit_type=iterative_build_omit omit_region_specification=omit_around_pdb omit_box_pdb=Mask.pdb\ omit_res_start_list="1 1" omit_res_end_list="39 30" omit_chain_list="Y Z" \ rebuild_in_place=Yes Mask.pdb is attached. Resolve is unable to create a mask. The log file, omit_box_id.log is also attached. Resolve creates a temp file named TMP.pdb which I would expect to contain the contents of my Mask.pdb. It only contains the first set of dummy atoms; chain Y. Then when Resolve reads this file it appears not to find even those atoms. I know Tom is out of reach for quite a while. Is there someone else who could help me over this hurdle? Dale Tronrud

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Autosol Feedback and questions
by Partha Chakrabarti 04 Jan '08

04 Jan '08

Happy new year everybody, I had quite a bit of difficulty with a SAD dataset which also showed Sulphur. Previously I had tried SHELEX to find sites, MLPHARE to refine, AddSOLVE to find more , and both SOLVE and PHASER to solve but it did not get to a point where I could do manual building. Autosol worked beautifully, the input files were scalepack merged data, ha.pdb (from addsolve step), and the sequence. PHASER to solve and TEXTAL & RESOLVE with through build option. This actually leave me with some confusions / questions: 1. When I compare the other "solutions", they look like the right one, just not good enough. Where exactly is the autosol making a difference? Is it the ability of of TEXTAL to build something more meaningful when the solution is not good or number of iterations? 2. How does PHASER know the different anomalous atoms? Does is use a different scaling factor for SE and S? 3. I see that phenix.refine has been used in the internal runs, I have used phenix.refine earlier. But after playing with the side chains in Coot, when I tried to use the modified pdb file and resolve.mtz, it complained about the good old Free_R flag. However, it ran directly in Refmac. Is it a known problem or maybe I did something stupid? I could do a dry run and change the read mtz and not cns if that would solve the problem. 4. Lastly, is it possible to use multiple processors to run the job? Best regards, Partha -- MRC National Institute for Medical Research Division of Molecular Structure The Ridgeway, NW7 1AA, UK Email: pchakra(a)nimr.mrc.ac.uk Phone: + 44 208 816 2515

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