[ccp4bb] Research Associate at Imperial College London
We wish to recruit a Research Associate to work in the research group of Professor Xiaodong Zhang (www.msf.bio.ic.ac.ukhttp://www.msf.bio.ac.uk), in the Section of Structural Biology, Department of Medicine, at Imperial College London’s South Kensington Campus. The successful candidate will join a multi-disciplinary team of international researchers investigating the structures and mechanisms of large key complexes in DNA damage response and repair. We employ a combination of X-ray crystallography and cryo-electron microscopy techniques, utilising our excellent structural biology facilities at Imperial. You must hold a PhD, or equivalent, in biochemistry, structural biology or relevant discipline and have a demonstrated track record in conducting high quality original research. For informal enquiries please contact Professor Xiaodong Zhang xiaodong.zh...@imperial.ac.ukmailto:xiaodong.zh...@imperial.ac.uk. Our preferred method of application is online via our website at http://www3.imperial.ac.uk/employment (please select “Job Search” then enter the vacancy reference number HM2014190 into “Keywords”). Please complete and upload an application form as directed quoting reference number HM2014190. Alternatively, if you are unable to apply online, please email hmr...@imperial.ac.ukmailto:hmr...@imperial.ac.uk to request an application form. The Closing date is 17 November, 2014 Professor Xiaodong Zhang Section of Structural Biology Department of Medicine Imperial College London 504, Sir Ernst Chain Building South Kensington, London, SW7 2AZ (0) 207 594 3151 xiaodong.zh...@imperial.ac.ukmailto:xiaodong.zh...@imperial.ac.uk http://www.imperial.ac.uk/people/xiaodong.zhang
Re: [ccp4bb] R free flag missing after ARP/wARP?
Dear Lu Zuokun, since you can use the output mtz-file from ArpWarp for model building, but not for refinement, there is no need to include the Rfree flag in its output file. Maybe the omission is a deliberate caveat to the users to pay attention to this. Best regards, Tim On 10/20/2014 04:41 AM, luzuok wrote: Dear all, I was using ARP/wARP in ccp4i, the input mtz file certainly has free R flag, but the output mzt file doesn't have the free R flag label? I chose do not use the Free R flag in the ARP/wARP GUI. Can anyone tell me what's wrong with this? best reagrds! Lu Zuokun -- 卢作焜 南开大学新生物站A202 -- Dr Tim Gruene Institut fuer anorganische Chemie Tammannstr. 4 D-37077 Goettingen GPG Key ID = A46BEE1A signature.asc Description: OpenPGP digital signature
[ccp4bb] protein forming dimers
Hello everyone, I would like to take a advice from you all. I have protein like 200 KDa when i lode in a gel it shows band on 200KDa, Is that this protein gets dimer or anything else is happening. I would appreciate for some open discussion. -- * Regards,** Arun *
Re: [ccp4bb] protein forming dimers
Well, If it is an SDS-PAGE gel, then the act of boiling the protein in detergent generally denatures the protein and breaks apart non-covalent interactions. To assess dimerisation, a non-denaturing technique might be more helpful. Native-PAGE, Size Exlusion Chromatography, Analytical Ultracentrifugation... SEC with a suitable column for your predicted protein size would probably be a first thing to try. If you have SEC with in-line MALLS, even better. HTH, Dave [image: David Briggs on about.me] David Briggs about.me/david_briggs http://about.me/david_briggs On 20 October 2014 12:00, arun kumar arungreenlo...@gmail.com wrote: Hello everyone, I would like to take a advice from you all. I have protein like 200 KDa when i lode in a gel it shows band on 200KDa, Is that this protein gets dimer or anything else is happening. I would appreciate for some open discussion. -- * Regards,** Arun *
Re: [ccp4bb] R free flag missing after ARP/wARP?
Dear Colin and Tim, But ARPwARP uses REFMAC5 for refinement, does this means that REFMAC5 uses all reflections as working set? How to validate that the refinement is not over fit? Best wishes! Lu Zuokun -- 卢作焜 南开大学新生物站A202 At 2014-10-20 17:38:58, Tim Gruene t...@shelx.uni-ac.gwdg.de wrote: Dear Lu Zuokun, since you can use the output mtz-file from ArpWarp for model building, but not for refinement, there is no need to include the Rfree flag in its output file. Maybe the omission is a deliberate caveat to the users to pay attention to this. Best regards, Tim On 10/20/2014 04:41 AM, luzuok wrote: Dear all, I was using ARP/wARP in ccp4i, the input mtz file certainly has free R flag, but the output mzt file doesn't have the free R flag label? I chose do not use the Free R flag in the ARP/wARP GUI. Can anyone tell me what's wrong with this? best reagrds! Lu Zuokun -- 卢作焜 南开大学新生物站A202 -- Dr Tim Gruene Institut fuer anorganische Chemie Tammannstr. 4 D-37077 Goettingen GPG Key ID = A46BEE1A
[ccp4bb] Fwd: mapsig errors
Hi all, I tried using mapsig for the first time, and I got multiple errors (text pasted at bottom). This is on mac os x 10.8.5, and the CCP4 install is 6.4.0. I followed these instructions to be able to use CCP4 programs at the command line: http://plested.wordpress.com/2012/10/31/run-ccp4-programs-from-the-command-line-on-mac-osx/ http://ccp4wiki.org/~ccp4wiki/wiki/index.php?title=How_to_run_CCP4_programs_from_the_command_line I have also tried the csh version as well, with the same results. Any advice would be greatly appreciated. This is my first time attempting to use CCP4 without the GUI, so if there's additional setup necessary, that would be great info to know. Thanks, Scott mapsig MAPIN H96LA28I_6pt5superI.pdb MAPIN2 H96LQ17I10_6pt5superI.pdb TYPE RATIO MAPOUT test.map BFONT COLOR=#FF!--SUMMARY_BEGIN-- pre ### ### ### ### CCP4 6.4: MAPSIG version 6.4 : ## ### User: horowsah Run date: 20/10/2014 Run time: 09:06:01 Please reference: Collaborative Computational Project, Number 4. 1994. The CCP4 Suite: Programs for Protein Crystallography. Acta Cryst. D50, 760-763. as well as any specific reference in the program write-up. !--SUMMARY_END--/FONT/B # # TRANSLATION FUNCTION MAP SIGNAL ANALYSIS. # # BE WARNED THAT THIS PROGRAM ASSUMES PERIODIC MAP BOUNDARIES, (I.E. THE LEFT-HAND EDGE ABUTS ONTO THE RIGHT-HAND EDGE) AND THEREFORE MAY NOT LOCATE PEAKS AT THE EDGES OF THE MAP CORRECTLY IN OTHER CASES. IN SUCH CASES THE MAP SHOULD BE EXPANDED IN ONE OR MORE DIRECTIONS TO MAKE IT PERIODIC IN ALL DIRECTIONS. BFONT COLOR=#FF!--SUMMARY_BEGIN-- html !-- CCP4 HTML LOGFILE -- hr !--SUMMARY_END--/FONT/B CCP4 library signal ccp4_map:No associated header (Error) raised in ccp4_cmap_open CCP4 library signal ccp4_map:Cannot open file (Error) raised in MRDHDR mapsig: Error in opening input map file. mapsig: Error in opening input map file. Times: User: 0.0s System:0.0s Elapsed: 0:00 /pre /html !--SUMMARY_END--/FONT/B -- Scott Horowitz, Ph.D. Research Associate Howard Hughes Medical Institute University of Michigan Department of Molecular, Cellular, and Developmental Biology Bardwell lab 830 N. University Ave, Room 4007 Ann Arbor, MI 48109 phone: 734-647-6683 fax: 734-615-4226 -- Scott Horowitz, Ph.D. Research Associate Howard Hughes Medical Institute University of Michigan Department of Molecular, Cellular, and Developmental Biology Bardwell lab 830 N. University Ave, Room 4007 Ann Arbor, MI 48109 phone: 734-647-6683 fax: 734-615-4226
Re: [ccp4bb] R free flag missing after ARP/wARP?
When you chose the option “Do not use the Free R flag”, then you were telling Refmac not to use cross-validation and therefore to use all reflections as the working set. My experience (doubtless very limited compared to the ARP/wARP developers) is that it’s significantly better to use the Rfree flag, which makes sense since you need cross-validation data in order for likelihood targets to be calibrated properly. But maybe the developers have some set of test cases that led them to choose the opposite as the default. Best wishes, Randy Read On 20 Oct 2014, at 14:24, luzuok luzuo...@126.com wrote: Dear Colin and Tim, But ARPwARP uses REFMAC5 for refinement, does this means that REFMAC5 uses all reflections as working set? How to validate that the refinement is not over fit? Best wishes! Lu Zuokun -- 卢作焜 南开大学新生物站A202 At 2014-10-20 17:38:58, Tim Gruene t...@shelx.uni-ac.gwdg.de wrote: Dear Lu Zuokun, since you can use the output mtz-file from ArpWarp for model building, but not for refinement, there is no need to include the Rfree flag in its output file. Maybe the omission is a deliberate caveat to the users to pay attention to this. Best regards, Tim On 10/20/2014 04:41 AM, luzuok wrote: Dear all, I was using ARP/wARP in ccp4i, the input mtz file certainly has free R flag, but the output mzt file doesn't have the free R flag label? I chose do not use the Free R flag in the ARP/wARP GUI. Can anyone tell me what's wrong with this? best reagrds! Lu Zuokun -- 卢作焜 南开大学新生物站A202 -- Dr Tim Gruene Institut fuer anorganische Chemie Tammannstr. 4 D-37077 Goettingen GPG Key ID = A46BEE1A -- 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 RoadE-mail: rj...@cam.ac.uk Cambridge CB2 0XY, U.K. www-structmed.cimr.cam.ac.uk
Re: [ccp4bb] mapsig errors
Please ignore my incredibly dumb last email… autocomplete had me putting in .pdb files, which obviously won't work. Scott On Mon, Oct 20, 2014 at 9:47 AM, Scott Horowitz horow...@umich.edu wrote: Hi all, I tried using mapsig for the first time, and I got multiple errors (text pasted at bottom). This is on mac os x 10.8.5, and the CCP4 install is 6.4.0. I followed these instructions to be able to use CCP4 programs at the command line: http://plested.wordpress.com/2012/10/31/run-ccp4-programs-from-the-command-line-on-mac-osx/ http://ccp4wiki.org/~ccp4wiki/wiki/index.php?title=How_to_run_CCP4_programs_from_the_command_line I have also tried the csh version as well, with the same results. Any advice would be greatly appreciated. This is my first time attempting to use CCP4 without the GUI, so if there's additional setup necessary, that would be great info to know. Thanks, Scott mapsig MAPIN H96LA28I_6pt5superI.pdb MAPIN2 H96LQ17I10_6pt5superI.pdb TYPE RATIO MAPOUT test.map BFONT COLOR=#FF!--SUMMARY_BEGIN-- pre ### ### ### ### CCP4 6.4: MAPSIG version 6.4 : ## ### User: horowsah Run date: 20/10/2014 Run time: 09:06:01 Please reference: Collaborative Computational Project, Number 4. 1994. The CCP4 Suite: Programs for Protein Crystallography. Acta Cryst. D50, 760-763. as well as any specific reference in the program write-up. !--SUMMARY_END--/FONT/B # # TRANSLATION FUNCTION MAP SIGNAL ANALYSIS. # # BE WARNED THAT THIS PROGRAM ASSUMES PERIODIC MAP BOUNDARIES, (I.E. THE LEFT-HAND EDGE ABUTS ONTO THE RIGHT-HAND EDGE) AND THEREFORE MAY NOT LOCATE PEAKS AT THE EDGES OF THE MAP CORRECTLY IN OTHER CASES. IN SUCH CASES THE MAP SHOULD BE EXPANDED IN ONE OR MORE DIRECTIONS TO MAKE IT PERIODIC IN ALL DIRECTIONS. BFONT COLOR=#FF!--SUMMARY_BEGIN-- html !-- CCP4 HTML LOGFILE -- hr !--SUMMARY_END--/FONT/B CCP4 library signal ccp4_map:No associated header (Error) raised in ccp4_cmap_open CCP4 library signal ccp4_map:Cannot open file (Error) raised in MRDHDR mapsig: Error in opening input map file. mapsig: Error in opening input map file. Times: User: 0.0s System:0.0s Elapsed: 0:00 /pre /html !--SUMMARY_END--/FONT/B -- Scott Horowitz, Ph.D. Research Associate Howard Hughes Medical Institute University of Michigan Department of Molecular, Cellular, and Developmental Biology Bardwell lab 830 N. University Ave, Room 4007 Ann Arbor, MI 48109 phone: 734-647-6683 fax: 734-615-4226 -- Scott Horowitz, Ph.D. Research Associate Howard Hughes Medical Institute University of Michigan Department of Molecular, Cellular, and Developmental Biology Bardwell lab 830 N. University Ave, Room 4007 Ann Arbor, MI 48109 phone: 734-647-6683 fax: 734-615-4226 -- Scott Horowitz, Ph.D. Research Associate Howard Hughes Medical Institute University of Michigan Department of Molecular, Cellular, and Developmental Biology Bardwell lab 830 N. University Ave, Room 4007 Ann Arbor, MI 48109 phone: 734-647-6683 fax: 734-615-4226
Re: [ccp4bb] R free flag missing after ARP/wARP?
Dear Lu Zuokun, The option you mention do not use the Free R flag, tells ARP/wARP to do refinement without using the R-free flags, so it does not seem strange to me that it would output a file without those labels. I don't know why this would be a good idea, though, if you want to continue building on this model with those same R-free flags. The manual says: The default is not to use Rfree, since the number of traced residues serves as excellent indicator of the success of the job. You can certainly turn the use of Rfree on. (http://www.embl-hamburg.de/ARP/Manual/UserGuide7.4.html) Maybe the thought is that the free reflections will be uncoupled during further building and refinement? Best regards, Folmer Fredslund 2014-10-20 4:41 GMT+02:00 luzuok luzuo...@126.com: Dear all, I was using ARP/wARP in ccp4i, the input mtz file certainly has free R flag, but the output mzt file doesn't have the free R flag label? I chose do not use the Free R flag in the ARP/wARP GUI. Can anyone tell me what's wrong with this? best reagrds! Lu Zuokun -- 卢作焜 南开大学新生物站A202 -- Folmer Fredslund
[ccp4bb] a more intelligent mapsig question
Hi all, Now that I have mapsig running, I am getting an error message that I am confused by. The maps I am using are anomalous maps generated by FFT. Originally, I had them covering just the asymmetric unit, but based on the error message below, I then redid it where I covered a user defined extent, where I went from 0 to the cell size for each axis, but the error message stayed the same, which means perhaps I'm misunderstanding what it means. Any help would be great, and the whole output is pasted below. Thanks, Scott bash-3.2$ mapsig mapin H96LY10I01_14combfull.map mapin2 H96LY10I01_6pt5combfull.map type ratio mapout test.map BFONT COLOR=#FF!--SUMMARY_BEGIN-- pre ### ### ### ### CCP4 6.4: MAPSIG version 6.4 : ## ### User: horowsah Run date: 20/10/2014 Run time: 10:20:01 Please reference: Collaborative Computational Project, Number 4. 1994. The CCP4 Suite: Programs for Protein Crystallography. Acta Cryst. D50, 760-763. as well as any specific reference in the program write-up. !--SUMMARY_END--/FONT/B # # TRANSLATION FUNCTION MAP SIGNAL ANALYSIS. # # BE WARNED THAT THIS PROGRAM ASSUMES PERIODIC MAP BOUNDARIES, (I.E. THE LEFT-HAND EDGE ABUTS ONTO THE RIGHT-HAND EDGE) AND THEREFORE MAY NOT LOCATE PEAKS AT THE EDGES OF THE MAP CORRECTLY IN OTHER CASES. IN SUCH CASES THE MAP SHOULD BE EXPANDED IN ONE OR MORE DIRECTIONS TO MAKE IT PERIODIC IN ALL DIRECTIONS. BFONT COLOR=#FF!--SUMMARY_BEGIN-- html !-- CCP4 HTML LOGFILE -- hr !--SUMMARY_END--/FONT/B Logical Name: MAPIN Filename: H96LY10I01_14combfull.map File name for input map file on unit 1 : H96LY10I01_14combfull.map file size 1917228 ; logical name MAPIN Number of columns, rows, sections ... 43 43 259 Map mode 2 Start and stop points on columns, rows, sections 0 420 420 258 Grid sampling on x, y, z 48 48 304 Cell dimensions . 42.7000 42.7000 258.299890.90.90. Fast, medium, slow axes .YXZ Minimum density .-0.04944 Maximum density . 0.19449 Mean density -0.0 Rms deviation from mean density . 0.01025 Space-group . 91 Number of titles 1 Labels: [No title given] Translation modulus = 43 43 259 *** ERROR - space group must be P1 if whole cell not given. -- Scott Horowitz, Ph.D. Research Associate Howard Hughes Medical Institute University of Michigan Department of Molecular, Cellular, and Developmental Biology Bardwell lab 830 N. University Ave, Room 4007 Ann Arbor, MI 48109 phone: 734-647-6683 fax: 734-615-4226
Re: [ccp4bb] Fwd: mapsig errors
Hi Scott, it's complaining that the input files are not in the standard CCP4 map format. Since they have the extension 'pdb' and not 'map' could they by any chance be PDB format files? If so then this won't work! Cheers -- Ian On 20 October 2014 14:47, Scott Horowitz horow...@umich.edu wrote: Hi all, I tried using mapsig for the first time, and I got multiple errors (text pasted at bottom). This is on mac os x 10.8.5, and the CCP4 install is 6.4.0. I followed these instructions to be able to use CCP4 programs at the command line: http://plested.wordpress.com/2012/10/31/run-ccp4-programs-from-the-command-line-on-mac-osx/ http://ccp4wiki.org/~ccp4wiki/wiki/index.php?title=How_to_run_CCP4_programs_from_the_command_line I have also tried the csh version as well, with the same results. Any advice would be greatly appreciated. This is my first time attempting to use CCP4 without the GUI, so if there's additional setup necessary, that would be great info to know. Thanks, Scott mapsig MAPIN H96LA28I_6pt5superI.pdb MAPIN2 H96LQ17I10_6pt5superI.pdb TYPE RATIO MAPOUT test.map BFONT COLOR=#FF!--SUMMARY_BEGIN-- pre ### ### ### ### CCP4 6.4: MAPSIG version 6.4 : ## ### User: horowsah Run date: 20/10/2014 Run time: 09:06:01 Please reference: Collaborative Computational Project, Number 4. 1994. The CCP4 Suite: Programs for Protein Crystallography. Acta Cryst. D50, 760-763. as well as any specific reference in the program write-up. !--SUMMARY_END--/FONT/B # # TRANSLATION FUNCTION MAP SIGNAL ANALYSIS. # # BE WARNED THAT THIS PROGRAM ASSUMES PERIODIC MAP BOUNDARIES, (I.E. THE LEFT-HAND EDGE ABUTS ONTO THE RIGHT-HAND EDGE) AND THEREFORE MAY NOT LOCATE PEAKS AT THE EDGES OF THE MAP CORRECTLY IN OTHER CASES. IN SUCH CASES THE MAP SHOULD BE EXPANDED IN ONE OR MORE DIRECTIONS TO MAKE IT PERIODIC IN ALL DIRECTIONS. BFONT COLOR=#FF!--SUMMARY_BEGIN-- html !-- CCP4 HTML LOGFILE -- hr !--SUMMARY_END--/FONT/B CCP4 library signal ccp4_map:No associated header (Error) raised in ccp4_cmap_open CCP4 library signal ccp4_map:Cannot open file (Error) raised in MRDHDR mapsig: Error in opening input map file. mapsig: Error in opening input map file. Times: User: 0.0s System:0.0s Elapsed: 0:00 /pre /html !--SUMMARY_END--/FONT/B -- Scott Horowitz, Ph.D. Research Associate Howard Hughes Medical Institute University of Michigan Department of Molecular, Cellular, and Developmental Biology Bardwell lab 830 N. University Ave, Room 4007 Ann Arbor, MI 48109 phone: 734-647-6683 fax: 734-615-4226 -- Scott Horowitz, Ph.D. Research Associate Howard Hughes Medical Institute University of Michigan Department of Molecular, Cellular, and Developmental Biology Bardwell lab 830 N. University Ave, Room 4007 Ann Arbor, MI 48109 phone: 734-647-6683 fax: 734-615-4226
Re: [ccp4bb] mapsig errors
Hi Scott Our mails crossed: you worked it out yourself a microsecond before I did! Cheers -- Ian On 20 October 2014 14:54, Scott Horowitz horow...@umich.edu wrote: Please ignore my incredibly dumb last email… autocomplete had me putting in .pdb files, which obviously won't work. Scott On Mon, Oct 20, 2014 at 9:47 AM, Scott Horowitz horow...@umich.edu wrote: Hi all, I tried using mapsig for the first time, and I got multiple errors (text pasted at bottom). This is on mac os x 10.8.5, and the CCP4 install is 6.4.0. I followed these instructions to be able to use CCP4 programs at the command line: http://plested.wordpress.com/2012/10/31/run-ccp4-programs-from-the-command-line-on-mac-osx/ http://ccp4wiki.org/~ccp4wiki/wiki/index.php?title=How_to_run_CCP4_programs_from_the_command_line I have also tried the csh version as well, with the same results. Any advice would be greatly appreciated. This is my first time attempting to use CCP4 without the GUI, so if there's additional setup necessary, that would be great info to know. Thanks, Scott mapsig MAPIN H96LA28I_6pt5superI.pdb MAPIN2 H96LQ17I10_6pt5superI.pdb TYPE RATIO MAPOUT test.map BFONT COLOR=#FF!--SUMMARY_BEGIN-- pre ### ### ### ### CCP4 6.4: MAPSIG version 6.4 : ## ### User: horowsah Run date: 20/10/2014 Run time: 09:06:01 Please reference: Collaborative Computational Project, Number 4. 1994. The CCP4 Suite: Programs for Protein Crystallography. Acta Cryst. D50, 760-763. as well as any specific reference in the program write-up. !--SUMMARY_END--/FONT/B # # TRANSLATION FUNCTION MAP SIGNAL ANALYSIS. # # BE WARNED THAT THIS PROGRAM ASSUMES PERIODIC MAP BOUNDARIES, (I.E. THE LEFT-HAND EDGE ABUTS ONTO THE RIGHT-HAND EDGE) AND THEREFORE MAY NOT LOCATE PEAKS AT THE EDGES OF THE MAP CORRECTLY IN OTHER CASES. IN SUCH CASES THE MAP SHOULD BE EXPANDED IN ONE OR MORE DIRECTIONS TO MAKE IT PERIODIC IN ALL DIRECTIONS. BFONT COLOR=#FF!--SUMMARY_BEGIN-- html !-- CCP4 HTML LOGFILE -- hr !--SUMMARY_END--/FONT/B CCP4 library signal ccp4_map:No associated header (Error) raised in ccp4_cmap_open CCP4 library signal ccp4_map:Cannot open file (Error) raised in MRDHDR mapsig: Error in opening input map file. mapsig: Error in opening input map file. Times: User: 0.0s System:0.0s Elapsed: 0:00 /pre /html !--SUMMARY_END--/FONT/B -- Scott Horowitz, Ph.D. Research Associate Howard Hughes Medical Institute University of Michigan Department of Molecular, Cellular, and Developmental Biology Bardwell lab 830 N. University Ave, Room 4007 Ann Arbor, MI 48109 phone: 734-647-6683 fax: 734-615-4226 -- Scott Horowitz, Ph.D. Research Associate Howard Hughes Medical Institute University of Michigan Department of Molecular, Cellular, and Developmental Biology Bardwell lab 830 N. University Ave, Room 4007 Ann Arbor, MI 48109 phone: 734-647-6683 fax: 734-615-4226 -- Scott Horowitz, Ph.D. Research Associate Howard Hughes Medical Institute University of Michigan Department of Molecular, Cellular, and Developmental Biology Bardwell lab 830 N. University Ave, Room 4007 Ann Arbor, MI 48109 phone: 734-647-6683 fax: 734-615-4226
Re: [ccp4bb] R free flag missing after ARP/wARP?
Dear Lu, The MTZ file produced at the end of the ARP/wARP protein model building is created by Refmac and contains the following: 1. New columns created for the built model: (FC, PHIC, FC_ALL, PHIC_ALL, FWT, PHWT, DELFWT, PHDELWT, FOM, FC_ALL_LS, PHIC_ALL_LS) 2. Columns from the input data file that were used for the refinement - structure factor amplitudes and their estimated standard deviations scaled to those calculated from the model. If Rfree flag was used, the FREE column is also copied to the output MTZ file. Since you chose not to use Rfree for model building, the Rfree flag was not copied to the output. Historically, the default for ARP/wARP protein model building is not to use Rfree. Some years ago we did observe a number of cases where setting a part of the data aside for cross-validation resulted in lower observation-to-parameter ratio and poorer model building. We also thought that the auto-traced model (if built) could itself serve as a validation criterion. In contrast, for ARP/wARP building the solvent structure only, the use of Rfree is a default. Over the recent years much has improved, notably Refmac's likelihood targets. I am sure that in many cases the use of Rfree could now be advantageous for protein model building. We will try to repeat extensive benchmarking with different ARP/wARP protocols, so that many default parameters could be improved. With best regards, Victor On 20/10/2014 04:41, luzuok wrote: Dear all, I was using ARP/wARP in ccp4i, the input mtz file certainly has free R flag, but the output mzt file doesn't have the free R flag label? I chose " do not use the Free R flag" in the ARP/wARP GUI. Can anyone tell me what's wrong with this? best reagrds! Lu Zuokun -- 卢作焜 南开大学新生物站A202
[ccp4bb] Normal mode refinement
Dear CCP4 Users, I seek your valuable advice and suggestion in carrying out the normal mode structure refinement which manifest the dynamics of protein as linear combination of harmonic modes, used to describe the motion of protein structure in collective fashion. Studies suggest that it is highly useful in refining the protein structure which harbors a considerable magnitude of flexibility in atomic position owing to high thermal factors. Therefor I want to know is there any software/script available to execute the normal mode of refinement. Thanks a lot in advance for your imperative suggestions Appu
Re: [ccp4bb] Normal mode refinement
On Monday, 20 October, 2014 18:10:03 Appu kumar wrote: Dear CCP4 Users, I seek your valuable advice and suggestion in carrying out the normal mode structure refinement which manifest the dynamics of protein as linear combination of harmonic modes, used to describe the motion of protein structure in collective fashion. Studies suggest that it is highly useful in refining the protein structure which harbors a considerable magnitude of flexibility in atomic position owing to high thermal factors. Therefor I want to know is there any software/script available to execute the normal mode of refinement. Thanks a lot in advance for your imperative suggestions The previously published examples of normal-mode refinement that I know about used private external programs to generate thermal ellipsoids for each atom, and then used those as fixed ADPs while refining coordinates in refmac or similar standard program. Again speaking only of the examples I have looked at in detail, the result was better (had lower R factors) than a conventional isotropic refinement but was not nearly as good as a multi-group TLS refinement of the same structure (TLSMD + refmac). On the other hand, there is a quite different way normal modes can be used in refinement. As I understand it (perhaps Garib will add addtional details) the jellybody refinement mode of recent refmac versions can be viewed as restraining the model shifts to be consistent with the principle normal mode. In this way the normal mode contributes to the path of the refinement, but is not explicitly part of the final model. So it may be that using TLSMD + refmac jellybody TLS refinement would get you the best of both approaches, though I have not gone back to look again at the published example structures since the advent of jellybody refinement. But note that jellybody is primarily useful when you already have a high-qualityl, good geometry, starting model. Ethan -- Ethan A Merritt Biomolecular Structure Center, K-428 Health Sciences Bldg MS 357742, University of Washington, Seattle 98195-7742
Re: [ccp4bb] Normal mode refinement
I agree with Ethan. In philosophy, NMA is a useful analysis to study low frequency collective motions. That is true by taking a stand-alone structure and explore such motions of biological interest. Domain motions in the crystallographic environment need not necessarily correspond to those of the isolated molecule. Also, please remember, many a times the most significant modes (say mode 7 or mode 8) in NMA do not represent the motions of biological interest. One is usually recommended to analyze many significant modes to extract the useful information, and in the extremes of argument, making at times this as a subjective exercise. In any case, I would strongly argue for TLSMD, as Ethan has pointed out, as that represents modeling disorder in crystallographic environment. In one of the examples where we were studying the relationship between NMA (for a stand-alone molecule) and TLS, we could correlate the two for the 11th and the 12th mode, suggesting clearly that 7th- 10th modes did not show a good correlation. By incorporating the 7-10th modes in crystallographic refinement, we would have clearly not made the model better! Shekhar On Tue, Oct 21, 2014 at 6:22 AM, Ethan A Merritt merr...@u.washington.edu wrote: On Monday, 20 October, 2014 18:10:03 Appu kumar wrote: Dear CCP4 Users, I seek your valuable advice and suggestion in carrying out the normal mode structure refinement which manifest the dynamics of protein as linear combination of harmonic modes, used to describe the motion of protein structure in collective fashion. Studies suggest that it is highly useful in refining the protein structure which harbors a considerable magnitude of flexibility in atomic position owing to high thermal factors. Therefor I want to know is there any software/script available to execute the normal mode of refinement. Thanks a lot in advance for your imperative suggestions The previously published examples of normal-mode refinement that I know about used private external programs to generate thermal ellipsoids for each atom, and then used those as fixed ADPs while refining coordinates in refmac or similar standard program. Again speaking only of the examples I have looked at in detail, the result was better (had lower R factors) than a conventional isotropic refinement but was not nearly as good as a multi-group TLS refinement of the same structure (TLSMD + refmac). On the other hand, there is a quite different way normal modes can be used in refinement. As I understand it (perhaps Garib will add addtional details) the jellybody refinement mode of recent refmac versions can be viewed as restraining the model shifts to be consistent with the principle normal mode. In this way the normal mode contributes to the path of the refinement, but is not explicitly part of the final model. So it may be that using TLSMD + refmac jellybody TLS refinement would get you the best of both approaches, though I have not gone back to look again at the published example structures since the advent of jellybody refinement. But note that jellybody is primarily useful when you already have a high-qualityl, good geometry, starting model. Ethan -- Ethan A Merritt Biomolecular Structure Center, K-428 Health Sciences Bldg MS 357742, University of Washington, Seattle 98195-7742 -- Shekhar C. Mande (शेखर चिं मांडे) Director, National Centre for Cell Science Ganeshkhind, Pune 411 007 Email: shek...@nccs.res.in, direc...@nccs.res.in Phone: +91-20-25708121 Fax:+91-20-25692259
Re: [ccp4bb] Normal mode refinement
Hello, You can also contact elNemo or NOMAD-Ref server developers about getting covariance/correlation matrices from normal mode analysis outputs to know the correctly coordinated mobile atoms. In this way you can compare with biological data also. In Shekhar's said paper K. Suhre (one of the developer of el-Nemo server) has done the same very correctly. best wishes, Arpita On Tue, Oct 21, 2014 at 5:40 AM, Appu kumar appu.kum...@gmail.com wrote: Dear CCP4 Users, I seek your valuable advice and suggestion in carrying out the normal mode structure refinement which manifest the dynamics of protein as linear combination of harmonic modes, used to describe the motion of protein structure in collective fashion. Studies suggest that it is highly useful in refining the protein structure which harbors a considerable magnitude of flexibility in atomic position owing to high thermal factors. Therefor I want to know is there any software/script available to execute the normal mode of refinement. Thanks a lot in advance for your imperative suggestions Appu -- Arpita -- Arpita Goswami Senior Research Fellow Structural Biology Laboratory Centre for DNA Fingerprinting and Diagnostics (CDFD) Tuljaguda (Opp MJ Market), Nampally, Hyderabad 500 001 INDIA Phone: +91- 40- 24749401/404 Mobile: 9390923667, 9502389184 Email: arp...@cdfd.org.in
