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Re: [phenixbb] what about heavy atom as model for MR
by zhaoy 29 Jun '13

29 Jun '13

Hi, It's my pleasure to recieved so much wonderful advice and I am trying according to the suggestions. I have used phenix.explore_metric_symmetry to see if there is any relationship between tetragonal and orthorhombic lattices. But the result suggests there is no relationship. In addition, I have some questions about the molecular replacement when density map entered as model. How to define clash between the model and How the value of RMS error of the density model impact the final MR result? Thank you! -- Yan Zhao, M.Phil. National Laboratory of Protein Sciences Institute of Biophysics Chinese Academy of Sciences 15 Datun Rd. Beijing, 100101 China Randy Read 写: Sorry, I'm coming a bit late to this thread, and most things I would have suggested have already been covered by Tom and Paul. Certainly there are examples in the literature of people using MR to place a heavy atom substructure against the anomalous differences, but as Tom pointed out there's the complication of making sure that you get the Se sites from one molecule and not from a combination of symmetry-related molecules, which probably won't have the same relative arrangement in the other crystal form. The one remaining thing I'd like to explore is whether you're sure that you've carried out the molecular replacement with density properly. This has become easier (with Tom's phenix.cut_out_density program), but it's still relatively complicated and error-prone. First, you have to be sure that you've cut out the density essentially for one molecule, and not a mixture of symmetry-related molecules. If the location and rough shape of the molecule isn't clear in your map, you may have to try things like cutting out spheres of density from different places in the unit cell and seeing which gives the clearest MR solution. Second, you have to make sure that you place the density in a reasonably large unit cell, 2.5-4 times the extent of the density that you've cut out, before using it as an MR model. Third, you have to choose an appropriate value for the effective RMS error of the density model. If your phases go to 4A, then a number like 1.5A to 2A might be appropriate. In our tests, the map can be pretty noisy but you can still get a very clear signal from the MR search. Of course, if the phase information is very weak, it still may not work even with an optimal envelope. Best wishes, Randy Read On 23 Jun 2013, at 15:56, "Terwilliger, Thomas C" <terwilliger(a)lanl.gov> wrote: You could try both the isomorphous and the anomalous differences. Each one has some useful information. You can even combine them but I'm guessing it won't help too much. Paul Adams' suggestion to check out related space groups is a very good one. Phenix.autosol only tries the opposite hand, so you have to try all the different screw axis possibilities that are not related by inversion. Another thing you could try is to repeat your MR with density from your map ( against the 2.6 A native) and get a long list of solutions. For each solution take the phases after MR and use them to calculate an anomalous difference Fourier for your semet P2221 data. Then notice if there are peaks in this map, and whether these peaks have the same relationship to the MR-placed density as you saw in your original map. If yes, you may have a solution. All the best, Tom T From: phenixbb-bounces(a)phenix-online.org [phenixbb-bounces(a)phenix-online.org] on behalf of zhaoy [zhaoy(a)moon.ibp.ac.cn] Sent: Sunday, June 23, 2013 3:32 PM To: PHENIX user mailing list Subject: Re: [phenixbb] what about heavy atom as model for MR Thank you very much. I will try as you said. The P2221 dataset's qulity are not very well. The points from diffraction image are not sharp. Besides, native dataset is 2.6A, setmet anomalous datasets is only 4A. So I have not obtained the solution with p2221datasets. There is only one molecular per AUS at p4322. So, from the initial map, I can confirm the heavy atom sites from the same molecular. Maybe I have a general idea about the method you said. But I still puzzled which F difference should I use, isomorphous or anomalous differences? Thanks! -- Yan Zhao, M.Phil. National Laboratory of Protein Sciences Institute of Biophysics Chinese Academy of Sciences 15 Datun Rd. Beijing, 100101 China Terwilliger, Thomas C 写: I'm not sure if there is a tool that will create the file you want for this. You could however just write a little script that reads in a .sca file and calculates F from I, subtract F+ - F- to get Dano, square that, write out Dano** 2 and sigma(Dano**2) as another .sca file. Then you could use this as input to molecular replacement with your sites from the P4322 dataset. There is an important caveat to this approach however: your sites from the P4322 have to be all part of the same molecule, and this might or might not happen automatically. If some sites are from one molecule and others from another molecule, then in the other crystal form their relationship won't be correct. If there is only one molecule in the au then this might or might not be a big problem. If there are more than one it is very likely to be a big problem. This would apply to the density search that you carried out as well. If your density is good enough to see where the molecule is, you might be able to figure out which of the symmetry-related sites to include in your MR search. Nevertheless as this is easy it seems like a fine thing to do. However I am wondering if it will work because your 2.6 A semet anomalous dataset did not yield a solution by itself, which is a little surprising... Also of course if your P4322 solution is not actually right or even just not close enough it would not work. -Tom T From: phenixbb-bounces(a)phenix-online.org [phenixbb-bounces(a)phenix-online.org] on behalf of 赵岩 [zhaoy(a)moon.ibp.ac.cn] Sent: Sunday, June 23, 2013 7:52 AM To: phenixbb(a)phenix-online.org Subject: [phenixbb] what about heavy atom as model for MR Hi everyone, The heavy atoms(Se-Met) were located at P4322 space group(4A). But the initial density map is so bad that I can not build the initial model. Meanwhile, I have another two datasets, native(2.6A) and se-met anomalous dataset with space group P2221. But these two datasets have no solution. I have tried the initial map from sp P4322 as a model to do molecular replacement. But no MR solution. So, I want to know how can I use the heavy atom as model to do MR with isomorphous or anomalous differences? Thanks! -- Yan Zhao, M.Phil. National Laboratory of Protein Sciences Institute of Biophysics Chinese Academy of Sciences 15 Datun Rd. Beijing, 100101 China _______________________________________________ phenixbb mailing list phenixbb(a)phenix-online.org http://phenix-online.org/mailman/listinfo/phenixbb ------ Randy J. Read Department of Haematology, University of Cambridge Cambridge Institute for Medical Research Tel: + 44 1223 336500 Wellcome Trust/MRC Building Fax: + 44 1223 336827 Hills Road E-mail: rjr27(a)cam.ac.uk Cambridge CB2 0XY, U.K. www-structmed.cimr.cam.ac.uk

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Is DEN refinement working?
by Wallen, Jamie 28 Jun '13

28 Jun '13

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PHASER-TFZ scores with ensembles higher?
by Jan Gebauer 28 Jun '13

28 Jun '13

Dear Phaser users, I recently tried to solve two difficult MR jobs by using ensembles in PHASER. With ENSEMBLES in this case I mean aligned structures prepared for PHASER by Phenix.ENSEMBLER. In both case I got solution with rather high TFZ-score (> 8), which I couldn't get wit individual structure, but was not able to refine them. My question is, whether using ensembles would increase the Z-Score without really improving phases. In other words, if there is another cut-off for the "Phaser solved it" Z-score, when using ensemble in contrast to single structures... Regards, Jan -- Dr. Jan Gebauer AG Prof. Baumann Institut für Biochemie / Uni-Köln Otto-Fischer-Str. 12-14 / 50674 Köln Fon: +49 (221) 470 3212 Fax: +49 (221) 470 5066 http://px.uni-koeln.de/

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refining the anomalous Se in MSE using Phenix Gui
by Kenneth A. Satyshur 27 Jun '13

27 Jun '13

Sirs: I am trying to refine the anomalous data in phenix refine. I am using the GUI and I get a box for the anomalous groups. But I when I put in 'Se' and refine: phenix says 'Sorry'. If I use View, I get all atoms in a viewer. I have 5 molecules of 330 residues each and 25 total Se atoms. I cannot possibly pick them out from the molecular display. Then I tried to make a heavy atom file with just selenium in it, but on input, it is read as a 'input model' and when I refine, it says 'duplicate atoms'. Without the anomalous refine, there is a lot of residual Fo-Fc density around the seleniums in the maps. How do I enter the 'anomalous group' into phenix refine. What is the syntax? I can find no help file or documentation on this. Anyone done this before? thanks -- Kenneth A. Satyshur, M.S.,Ph.D. Senior Scientist University of Wisconsin Madison, Wisconsin 53706 608-215-5207

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Phenix version 1.8.2 released
by Paul Adams 26 Jun '13

26 Jun '13

The Phenix developers are pleased to announce that version 1.8.2 of Phenix is now available. Binary installers for Linux, Mac OSX, and Windows platforms are available at the download site: http://phenix-online.org/download/ Highlights from this version: Ensemble refinement, updated hydrogen parameters in refinement and validation, mmCIF reading and writing, LLG maps output from refinement, ligand pipeline for automated solution of protein/ligand complexes, autobuilding starting from a very accurate but very small part of a model, CC* calculation from unmerged data, maximum entropy maps, and PDB editor. Graphical interface: ==================== - New GUIs: phenix.merging_statistics, phenix.pdb_editor General: ======== - Multiple bug fixes - phenix.refine, phenix.reduce, and MolProbity updated to use consistent, revised, electron-cloud centers for hydrogen positions as default (for X-ray) - also see MolProbity website for details: http://molprobity.biochem.duke.edu - mmCIF format now supported in place of PDB format for input to many programs - new commands: phenix.ensemble_refinement, phenix.ligand_pipeline, phenix.sort_hetatms, phenix.cc_star, phenix.pdb_editor, phenix.maximum_entropy_map Refinement: =========== - phenix.refine - support for LLG map type - uses Phaser to compute residual anomalous LLG map - best when used with group_anomalous strategy, which will flatten the LLG map around existing anomalous scatterers with refined f'/f''. - alternatively, anomalous_residual map type (similar but less sensitive) - automatic linking in refinement and pdbtools for residues in the same chain - enabled with automatic_linking.intra_chain=True; this includes carbohydrate linking to protein and between sugars, and covalently bound ligands. - optional output of mmCIF format model and data files - phenix.ensemble_refinement (new command): - New method for refining ensemble models - Combines MD simulation with X-ray restraints that simultaneously account for anisotopic and anharmonic atomic distributions. - For full description see: Burnley BT, Afonine PV, Adams PD, Gros P. 2012. Modelling dynamics in protein crystal structures by ensemble refinement. eLife 1:e00311. doi: 10.7554/elife.00311. Experimental Phasing and Model Building: ======================================== - phenix.autobuild: - now uses torsion NCS parameterization when NCS is used in refinement - building starting from a very accurate but very small part of a model: You can now use the keyword rebuild_from_fragments=True to start rebuilding from fragments of a model. You might want to use this if you look for ideal helices using Phaser, then rebuild the resulting partial model, as in the Arcimboldo procedure. The special feature of finding helices is that they can be very accurately placed in some cases. This really helps the subsequent rebuilding. If you have enough computer time, then run it several or even many times with different values of i_ran_seed. Each time you'll get a slightly different result. - Base-pairing in RNA building -- phenix.autobuild will now try to guess which bases in a model are base-paired, and if there is no positive sequence match to the model, the bases that are base-paired will be chosen to be complementary. You can set the cutoff for base pairing with the keyword dist_cut_base. - waters are automatically named with the chain of the closest macromolecule if you set sort_hetatms=True. This is for the final model only. - you can supply a target position for your model with map_to_object=my_target.pdb. Then at the very end of the run your molecule will be placed as close to this as possible. The center of mass of the autobuild model will be superimposed on the center of mass of my_target.pdb using space group symmetry, taking any match closer than 15 A within 3 unit cells of the original position. The new file will be overall_best_mapped.pdb. Molecular Replacement: ====================== - phenix.mr_rosetta - You can now give commands for Rosetta in mr_rosetta, including a command to specify where disulfide bonds are located - The Rosetta models are now identified by an ID number so that they have unique names - Default number of homology models to download is now 1 (was 5) - Default number of NCS copies (if ncs_copies=Auto) is number leading to solvent content closest to 50%; if ncs_copies=None then all plausible values of ncs_copies are tested. New commands: ============= - phenix.ligand_pipeline (new command): - combines Xtriage, Phaser, eLBOW, phenix.refine, AutoBuild, and LigandFit to automatically solve protein/ligand complexes - optional integration with Coot (parameter interactive=True) allows semi-interactive operation where more rebuilding is required - phenix.cc_star (new command): - combined assessment of model and data quality, as outlined in Karplus & Diederichs (2012) Science 336:1030-3. - summary also output by phenix.model_vs_data if additional unmerged_data keyword is specified - phenix.maximum_entropy_map (new command): - tool to compute maximum entropy map from map coefficients. The program reads input Fourier map coefficients and modifies corresponding synthesis using Maximum-Entropy Method (MEM). The MEM modified map is everywhere positive, smooth and is of higher resolution. The method uses is a modification of Gull & Daniell (1978) algorithm. - phenix.sort_hetatms (new command): - rearranges heteroatoms (ligands, waters, etc.) in a model so they are paired with the nearest macromolecule chain, similar to the PDB's processing of models - phenix.pdb_editor (new command): - graphical editor for PDB files, based on tree view of model hierarchy - perform common operations such as chain renaming, renumbering, manipulation of atomic properties, add/remove atoms Ligands: ======== - phenix.ready_set: - methyl rotations now automatic when adding explicit hydrogens (also in phenix.reduce) - more options adding deuteriums to a model have been introduced Miscellaneous: ============== - phenix.xtriage: - include merging statistics if unmerged intensities are used as input - phenix.maps: - added wavelength parameter and support for anomalous residual and Phaser LLG map coefficients For a full list of changes see: http://www.phenix-online.org/documentation/CHANGES Please note that this publication should be used to cite use of Phenix: PHENIX: a comprehensive Python-based system for macromolecular structure solution. P. D. Adams, P. V. Afonine, G. Bunkóczi, V. B. Chen, I. W. Davis, N. Echols, J. J. Headd, L.-W. Hung, G. J. Kapral, R. W. Grosse-Kunstleve, A. J. McCoy, N. W. Moriarty, R. Oeffner, R. J. Read, D. C. Richardson, J. S. Richardson, T. C. Terwilliger and P. H. Zwart. Acta Cryst. D66, 213-221 (2010). Full documentation is available here: http://www.phenix-online.org/documentation/ There is a Phenix bulletin board: http://www.phenix-online.org/mailman/listinfo/phenixbb/ Please consult the installer README file or online documentation for installation instructions. Direct questions and problem reports to the bulletin board or: help(a)phenix-online.org and bugs(a)phenix-online.org Commercial users interested in obtaining access to Phenix should visit the Phenix website for information about the Phenix Industrial Consortium. The development of Phenix is principally funded by the National Institute of General Medical Sciences (NIH) under grant P01-GM063210. We also acknowledge the generous support of the members of the Phenix Industrial Consortium. -- Paul Adams Deputy Division Director, Physical Biosciences Division, Lawrence Berkeley Lab Division Deputy for Biosciences, Advanced Light Source, Lawrence Berkeley Lab Adjunct Professor, Department of Bioengineering, U.C. Berkeley Vice President for Technology, the Joint BioEnergy Institute Laboratory Research Manager, ENIGMA Science Focus Area Building 64, Room 248 Tel: 1-510-486-4225, Fax: 1-510-486-5909 http://cci.lbl.gov/paul Lawrence Berkeley Laboratory 1 Cyclotron Road BLDG 64R0121 Berkeley, CA 94720, USA. Executive Assistant: Louise Benvenue [ LBenvenue(a)lbl.gov ][ 1-510-495-2506 ] --

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what about heavy atom as model for MR
by 赵岩 26 Jun '13

26 Jun '13

Hi everyone, The heavy atoms(Se-Met) were located at P4322 space group(4A). But the initial density map is so bad that I can not build the initial model. Meanwhile, I have another two datasets, native(2.6A) and se-met anomalous dataset with space group P2221. But these two datasets have no solution. I have tried the initial map from sp P4322 as a model to do molecular replacement. But no MR solution. So, I want to know how can I use the heavy atom as model to do MR with isomorphous or anomalous differences? Thanks! -- Yan Zhao, M.Phil. National Laboratory of Protein Sciences Institute of Biophysics Chinese Academy of Sciences 15 Datun Rd. Beijing, 100101 China

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possible Xtriage bug??
by Michael Thompson 25 Jun '13

25 Jun '13

Hello Developers, I would like to report a possible bug in newer versions of phenix.xtriage. I have noticed recently that I am getting some strange results for the results of the L-test when I run xtriage, as described below: My data show a strong non-crystallographic translation, described by the vector <uvw>=(1/3, 2/3, 1/2). This NCS translation is readily identified by xtriage. When Xtriage performs the L-test, the output states: L test for acentric data using difference vectors (dh,dk,dl) of the form: (2hp,2kp,2lp) where hp, kp, and lp are random signed integers such that 2 <= |dh| + |dk| + |dl| <= 8 Mean |L| :0.504 (untwinned: 0.500; perfect twin: 0.375) Mean L^2 :0.340 (untwinned: 0.333; perfect twin: 0.200) These results don't make sense, as the statistics show an "anti-twinned" effect, which should not happen with the L-test. It appears that xtriage is using reflections that do not have the same expected intensity values due to the NCS translation. The strange thing is that I have another xtriage logfile, which I produced in September of 2012 using the same input data, that indicates the reflections used for the L-test were chosen "using difference vectors (dh,dk,dl) of the form: (3hp,3kp,2lp)." This choice of reflections does account for proper parity, and I get results that are much more sensible (Mean |L|=0.394; Mean L^2=0.221). I am wondering why I am getting different results now than I did last September? Has something changed in the underlying code, or am I missing something silly? I do not have an easy way of determining which version of phenix I used when the L-test was conducted using sensible reflections (the xtriage logfile doesn't contain this info as far as I can tell), but the version that seems to choose reflections inappropriately is 1.8.2-1334. Thanks a lot for the help, and for phenix being awesome in general! Mike -- Michael C. Thompson Graduate Student Biochemistry & Molecular Biology Division Department of Chemistry & Biochemistry University of California, Los Angeles miket(a)chem.ucla.edu

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How to treat EDTA on a special position
by Jan Gebauer 25 Jun '13

25 Jun '13

Dear all, I recently solved a structure in R32/H (155). Interestingly some crystal contacts are mediated by an Metal/EDTA complex. The metal -presumably a magnesium- sits on a special position (x,0, 1/2) in the middle of a twofold axis with the EDTA molecule coordinating it. (See "top" view in dropbox image below) If I fit the EDTA/Mg complex in the density, the symmetry will of course generate a symmetry mate on the same spot. (see "side" view in dropbox image below). In phenix.refine, both entities are refined, which of course gives wrongs results. My question is, how to best define the Ligand to be compatible with phenix.refine and -later- with pdb submission. Should I just set occupancy to 0.5 for the whole EDTA molecule? - But then I would still get two molecules at this site... Or split it somehow half and let the second half be generated by symmetry? In that case, I would need to define bonds over the asymmetric unit to prevent "dissociation" of my ligand halves, wouldn't I? Or is there a third way? I already searched the phenixBB, but only found some "old" threads discussing vaguely similar problems. Unfortunately, they didn't help me to answer which way is better... (or right...) Regards, Jan DROPBOX IMAGE: https://dl.dropboxusercontent.com/u/29745394/R32%20-%20EDTA.jpg -- Dr. Jan Gebauer AG Prof. Baumann Institut für Biochemie / Uni-Köln Otto-Fischer-Str. 12-14 / 50674 Köln Fon: +49 (221) 470 3212 Fax: +49 (221) 470 5066 http://px.uni-koeln.de/

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completeness in table1
by Alexander Batyuk 24 Jun '13

24 Jun '13

Dear developers, What could be the reason for discrepancy in completeness reported in XSCALE.LP (94.5% - 100.0%) vs from Table1: 89.57% (37.49%)? Best wishes, Alex -- Alex Batyuk The Plueckthun Lab www.bioc.uzh.ch/plueckthun

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Re: [phenixbb] How to export difference patterson map in PHENIX
by Ulrich Baumann 20 Jun '13

20 Jun '13

Hi Nat I tried to limit the resolution by either --diff_limit 4 or --high_resolution 4.0. This does not work - probably is not supposed to work in this way? Thanks, ULrich lrich-Baumanns-iMac:xds_new ubaumann$ cctbx.patterson_map au1_run1_Rfree.mtz --high_resolution 4. !!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!! !! WARNING - EXPERIMENTAL PROGRAM !! !! !! !! This program is still in development - some functionality may be !! !! missing and/or untested. Use at your own risk! For bug reports, etc. !! !! email bugs(a)phenix-online.org. !! !!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!!! Traceback (most recent call last): File "/usr/local/phenix-dev-1405/build/mac-intel-osx-x86_64/../../cctbx_project/iotbx/command_line/patterson_map.py", line 176, in <module> run(sys.argv[1:]) File "/usr/local/phenix-dev-1405/build/mac-intel-osx-x86_64/../../cctbx_project/iotbx/command_line/patterson_map.py", line 136, in run params = cmdline.work.extract() File "/usr/local/phenix-dev-1405/cctbx_project/libtbx/phil/__init__.py", line 1690, in extract value = object.extract(parent=result) File "/usr/local/phenix-dev-1405/cctbx_project/libtbx/phil/__init__.py", line 1086, in extract return self._type_from_words()(self.words, master=self) File "/usr/local/phenix-dev-1405/cctbx_project/libtbx/phil/__init__.py", line 320, in from_words value = self._value_from_words(words=words, path=master.full_path()) File "/usr/local/phenix-dev-1405/cctbx_project/libtbx/phil/__init__.py", line 348, in _value_from_words return float_from_words(words=words, path=path) File "/usr/local/phenix-dev-1405/cctbx_project/libtbx/phil/__init__.py", line 274, in float_from_words result = number_from_words(words=words, path=path) File "/usr/local/phenix-dev-1405/cctbx_project/libtbx/phil/__init__.py", line 227, in number_from_words value_string=str_from_words(words), words=words, path=path) File "/usr/local/phenix-dev-1405/cctbx_project/libtbx/phil/__init__.py", line 212, in number_from_value_string path, value_string, words[0].where_str())) RuntimeError: Error interpreting high_resolution="True" as a numeric expression (command line argument, line 1) Ulrich-Baumanns-iMac:xds_new ubaumann$ cctbx.patterson_map au1_run1_Rfree.mtz --diff_limit 4 ---------------------------------------- Prof. Ulrich Baumann, PhD Institute of Biochemistry, University of Cologne Otto-Fischer-Strasse 12-14, D-50674 Cologne, Germany

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