[ccp4bb] AW: [ccp4bb] ANISOU in pdb and density improvement
Dear Appu, It seems that the anisotropy server you used put the anisotropic corrections in the pdb, which got subsequently rejected by the refinement programs because of insufficient resolution. The alternative is to apply anisotropic scaling to your reflection data, which is e.g. done by the UCLA anisotropy server. Using the scaled mtz should solve your problem. Best, Herman Von: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] Im Auftrag von Appu kumar Gesendet: Donnerstag, 2. Juli 2015 05:17 An: CCP4BB@JISCMAIL.AC.UK Betreff: Re: [ccp4bb] ANISOU in pdb and density improvement Hello All, Sorry for giving incomplete information. The protein is 75KDa membrane protein and it exists as tetramer in ASU. Structure is solved by MR. Overall completeness of data is 98% wiith multiplicity of 4.8. Density looks great after refinement having ANISOU record in PDB. Any suggestions and guidance will be much appreciated Thanks you Appu On 1 July 2015 at 21:14, xaravich ivan xaravich.i...@gmail.commailto:xaravich.i...@gmail.com wrote: 4 Angstrom resolution is pretty low and there has to be other info associated with that to get more help from here. How big is your protein? How are you solving the phases? How complete is your data at that resolution? What kind of multiplicity are you getting? I think you have other issues that are much more pressing than what you are asking here!!! As you are able to get 4A data it is evident that you can crystallize your protein. Congrats Now, concentrate on that and try to get higher resolution data from better diffracting crystals and it might be a much easier experience, (unless you trying to solve a huge ribosomal complex kind of protein and in that case 4 angs is amazing) cheers Ivan On Wed, Jul 1, 2015 at 5:57 PM, Appu kumar appu.kum...@gmail.commailto:appu.kum...@gmail.com wrote: Dear CCP4 users, I am refining a structure at 4A resolution. Crystal diffracted anisotropically and after refinement in both PHENIX and REFMAC, electron density in one of the domain of protein is represented by discontinuity and poor maps. Therefore i did the aniosotropy correction using anisotropy server. After anisotropy correction using anisotropy server, the electron density becomes resonable for 4A map as evident by appearance of electron density around some bulky sidechains and discontinuity in electron density is diminished. When i look into the pdb, I found that pdb contain ANISOU records for each atoms. I tried same things with other data sets and same data set again (data set described above) of same protein. After refinement, the ANISOU records are not present for each atoms in refined pdb and the electron density is discontinuous and poor. I am wondering why one of the anisotropically corrected MTZ by diffraction server gives the ANISOU records with the good density after refinement and other mtz file do not produce ANISOU record in pdb and poor density map. I would appreciate your help regarding this issue. Thank you Appu
[ccp4bb] Problem with ViewHKL 1.08
Dear all; I have reinstalled ccp4i recently and everything seems to work fine but ViewHKL. Every time I try to visualize a .mtz file the program gets opened but without loading the .mtz. Then it gets frozen and I have to kill it. In the command line the messages I get are: **SYMMETRY OPERATOR ERROR** Error in interpreting symop -X+1/2, -Y, Z+1/2 CCP4 library signal ccp4_parser:Failed to interpret symop string (Success) raised in symop_to_mat4 **SYMMETRY OPERATOR ERROR** Error in interpreting symop X+1/2, -Y+1/2, -Z CCP4 library signal ccp4_parser:Failed to interpret symop string (Success) raised in symop_to_mat4 **SYMMETRY OPERATOR ERROR** Error in interpreting symop -X, Y+1/2, -Z+1/2 CCP4 library signal ccp4_parser:Failed to interpret symop string (Success) raised in symop_to_mat4 Spacegroup_registry: ASU warning, LGhash=0x33b45b40 Spacegroup_registry: ASU fail File /media/marta/TOSHIBA/Ciencia/17_koji/20150626Xaloc/ch186/x4/b2x4/ccp4irepro/b2x4_3_0001_scaled1_refmac1.mtz is not an MTZ file My operative system is Linux Mint 17 (Qiana). Does some of you have any idea of what may be happening and how I can solve it? Thank you very much, Best regards Marta Sanz
[ccp4bb] WG: [ccp4bb] Real space refinement of alternative conformations
For the benefit of the whole bullitin board… Von: Schreuder, Herman RD/DE Gesendet: Donnerstag, 2. Juli 2015 09:26 An: 'Isaac Westwood' Betreff: AW: [ccp4bb] Real space refinement of alternative conformations Dear Isaac, I have done some tests and you are right, the peptide get seriously distorted. I was never able to do a zone real space refinement including alternate conformations in coot, but at least it was possible to refine one of the alternate conformations on its own. Apparently, Paul Emsley has closed this loophole in the newer versions of coot. I also tried real space refinement of 2 or 3 alternate residue in a row, which used to be ok as long as only one of the alternative zones was picked. Here I noticed some interesting behavior: The first and last (or only last in case of 2 residues) alternate residues refined correctly, and the second residue did not move at all. So an (ugly) workaround might be to define another alternative residue N-terminal from the residue you want to refine, refine the two and delete the dummy alternative residue afterwards. This might be less work than deleting and re-adding the alternative conformations. However, I believe that Paul should really fix this in a next version of coot. Best, Herman Von: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] Im Auftrag von Isaac Westwood Gesendet: Mittwoch, 1. Juli 2015 18:22 An: CCP4BB@JISCMAIL.AC.UKmailto:CCP4BB@JISCMAIL.AC.UK Betreff: [ccp4bb] Real space refinement of alternative conformations This is a cross-post from the cootbb last week, regarding the modelling of multiple conformations in coot. I know this has long been a tricky thing to do, and in the past I've always managed to model multiple alternative conformers by hook or by crook, but in some current structures I'm having a bit of a nightmare getting things to refine with reasonable geometry, and was wondering if anyone has noticed this and/or managed to resolve it satisfactorily. If you add an alternate conformer in coot, you cannot then do real space refinement with either sphere refine or real space refine zone (eg i +/- 1 by pressing 'a' or by selecting a zone around the residue with multiple conformers). In the case of sphere refine, the i-1 backbone peptide will become non-planar (or completely flip so the C=O bond overlaps the C-N bond), and the i peptide will also be distorted though to a lesser extent. In the case of real space refine zone, the i-1 and i backbone angles are distorted such that C-N-CA is around 150 degrees. You can real space refine zone, then just select one of the multiple conformers, which has worked acceptably in the past (especially when then fed into a refinement package, which tends to clear things up), but at least on the several datasets I've tested it on, this is still leading to distorted geometry, particularly peptide planarity of the i-1 and i peptides. I'd really like to get a model with good geometry directly from coot, so I don't have to rely on maximum likelihood refinement to sort out the problems - I always find that a rather clunky solution. Has anyone else noted or documented this behaviour, and is it possible to fix this? I know I may be asking a lot, here! To clarify some of the obvious questions up front: I'm using coot 0.8.1 on Ubuntu 12.04 and coot 0.8 on Max OSX with the same results. The observed behaviour occurs with and without the following restraints in coot: planar peptides, torsion and Ramachandran restraints, with a refinement weight typically of about 10, but I've observed the same problem at any value between 0.1 and 90 (I haven't tested wider as there's not much point!). You don't need high res data with genuine alt conf density to test this, you get the same behaviour by modelling in 2 x 0.5 occ conformers into the same density. The only work around so far seems to be real-space refining without the second conformer to get all the geometries right, then putting in the alt conf and selecting the nearest rotamer, then do ML refinement to try to fix the errors and get B-factors. Deleting the alt conf in coot before any additional real space refinement is then required (but coot cleverly remembers the B factors for the alt conf when you add it back in, which is neat). I'd be extremely grateful for any help to resolve this! Thanks in advance Isaac Westwood
[ccp4bb] AW: [ccp4bb] Rfree below Rwork
You are right. After I sent the email to the bulletin board, I realized that in R32 there must be more then unit cells but did not send a correction. Next time, I will check the space group before sending an email. Best regards, Herman Von: Oganesyan, Vaheh [mailto:oganesy...@medimmune.com] Gesendet: Donnerstag, 2. Juli 2015 15:48 An: Schreuder, Herman RD/DE; CCP4BB@JISCMAIL.AC.UK Betreff: RE: [ccp4bb] Rfree below Rwork Hi Herman, While you're correct regarding increase in number of entities in the asu upon lowering the symmetry, you're not correct for specific case of R32. One molecule per asu in R32 equals 18 molecules per asu in P1. Regards, Vaheh Oganesyan www.medimmune.com From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of herman.schreu...@sanofi.commailto:herman.schreu...@sanofi.com Sent: Wednesday, July 01, 2015 7:34 AM To: CCP4BB@JISCMAIL.AC.UKmailto:CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] AW: [ccp4bb] Rfree below Rwork Dear Boaz, One can equally well describe a R32 crystal with one molecule in the asymmetric unit as P1 and 6 molecules in the asymmetric unit. In this case, the NCS in P1 is identical to the crystallographic symmetry in R32. Best, Herman Von: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] Im Auftrag von Boaz Shaanan Gesendet: Mittwoch, 1. Juli 2015 12:10 An: CCP4BB@JISCMAIL.AC.UKmailto:CCP4BB@JISCMAIL.AC.UK Betreff: Re: [ccp4bb] Rfree below Rwork Just wondering about Eleanor's interesting remark: would the Rf Rw go as low as reported by Wolfram (0.22) in case of a wrong space group? Boaz Boaz Shaanan, Ph.D. Dept. of Life Sciences Ben-Gurion University of the Negev Beer-Sheva 84105 Israel E-mail: bshaa...@bgu.ac.ilmailto:bshaa...@bgu.ac.il Phone: 972-8-647-2220 Skype: boaz.shaanan Fax: 972-8-647-2992 or 972-8-646-1710 From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] on behalf of Eleanor Dodson [eleanor.dod...@york.ac.uk] Sent: Tuesday, June 30, 2015 8:55 PM To: CCP4BB@JISCMAIL.AC.UKmailto:CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] Rfree below Rwork I suppose if I was the referee for this structure and your FreeR is so close to the Rfactor I would ask you to ensure you had the right space group - is the 6 fold NCS actually 2 fold NCS with a crystallographic 3 fold.. Cases occur where R32 is indexed as C2.. Certainly if the Rfree set is assigned randomly to reflections which are symmetry equivalents then you see this phenomena of Rfree = Rfactor Eleanor On 30 June 2015 at 18:26, Gerard Bricogne g...@globalphasing.commailto:g...@globalphasing.com wrote: Dear Wolfram, I have a perhaps optimistic view of the effect of high-order NCS on Rfree, in the sense that I don't view it as a problem. People have agonised to extreme degrees over the difficulty of choosing a free set of reflections that would produce the expected gap between Rwork and Rfree, and some of the conclusions were that you would need to hide almost half of your data in some cases! I think it is best to remember that the idea of cross-validation by Rfree is to prevent overfitting, i.e. ending up with a model that fits the amplitudes too well compared to how well it determines the phases. In the case of high-order NCS (in your case, the U/V ratio that the old papers on NCS identified as the key quantity to measure the phasing power of NCS would be less than 0.1!) the phases and the amplitudes are so tightly coupled that it is simply impossible to fit the amplitudes without delivering phases of an equally good quality. In other words there is no overfitting problem (provided you do have good and complete data) and the difference between Rfree and Rwork is simply within the bounds of the statistical spread of Rfree depending on the free set chosen. You are lucky to have 6-fold NCS, so don't let any reviewer convince you that it is a curse, and make you suffer for it :-) . With best wishes, Gerard. -- On Tue, Jun 30, 2015 at 12:58:44PM -0400, wtempel wrote: Hello, my question concerns refinement of a structure with 6-fold NCS (local automatic restraints in REFMAC) against 2.8 A data. The size of my free set is 1172 selected in thin resolution shells (SFTOOLS) and corresponding to 4.3 % of reflections. A refmac run of 10 cycles of TLS and 10 cycles of CGMAT starts out at Rfree/Rcryst 0.271/0.272. After the 10th TLS cycle I have 0.227/0.224. Yes, Rfree Rcryst. At the end of CGMAT I have 0.2072/0.2071. I understand that NCS stresses the independence assumption of the free set. Am I correct in believing that Rfree *may* be smaller than Rcryst even in the absence of a major mistake? My hope is that the combined wisdom of ccp4bb followers can point out my possible mistake, suggest tests that I may perform to avoid them and, possibly, arguments in defense of a crystallographic model with Rfree Rcryst. Many thanks, Wolfram Tempel --
[ccp4bb] accept MR solution even if the number of clashes is above the threshold
Hi everyone, I am running Phaser (from Phenix) and while checking the .log file (it is still running) I realize that it found some solutions with a TFZ score over 7, but it won't take them I guess because the number of clashes is higher than allowed (12, I guess they're not so many either). My models are poor and for so I would like to check these solutions that for now Phaser will reject. However in the input I don't want to increase the allowed number of clashes because this will then affect all the other solutions and this will take forever... So my question is: can I tell phaser somehow to write down/save the solutions for which the TFZ score will be higher than 7? Even though the clashes Thanks a lot in advance All the best, Almu -- Almudena Ponce-Salvatierra Macromolecular crystallography and Nucleic acid chemistry Max Planck Institute for Biophysical Chemistry Am Fassberg 11 37077 Göttingen Germany
Re: [ccp4bb] AW: [ccp4bb] Rfree below Rwork
Hi Herman and Boaz, in the trigonal setting R32 (not in the hexagonal setting H32), the unit cell in R32 contains 6 copies. If you take the whole R32 unit cell as a P1 cell, you would have 6 copies in the asymmetric unit, as Hermann wrote. Best regards, Dirk. Am 02.07.15 um 15:52 schrieb herman.schreu...@sanofi.com: You are right. After I sent the email to the bulletin board, I realized that in R32 there must be more then unit cells but did not send a correction. Next time, I will check the space group before sending an email. Best regards, Herman *Von:*Oganesyan, Vaheh [mailto:oganesy...@medimmune.com] *Gesendet:* Donnerstag, 2. Juli 2015 15:48 *An:* Schreuder, Herman RD/DE; CCP4BB@JISCMAIL.AC.UK *Betreff:* RE: [ccp4bb] Rfree below Rwork Hi Herman, While you’re correct regarding increase in number of entities in the asu upon lowering the symmetry, you’re not correct for specific case of R32. One molecule per asu in R32 equals 18 molecules per asu in P1. /Regards,/ // /Vaheh Oganesyan/ /www.medimmune.com/ *From:*CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] *On Behalf Of *herman.schreu...@sanofi.com mailto:herman.schreu...@sanofi.com *Sent:* Wednesday, July 01, 2015 7:34 AM *To:* CCP4BB@JISCMAIL.AC.UK mailto:CCP4BB@JISCMAIL.AC.UK *Subject:* [ccp4bb] AW: [ccp4bb] Rfree below Rwork Dear Boaz, One can equally well describe a R32 crystal with one molecule in the asymmetric unit as P1 and 6 molecules in the asymmetric unit. In this case, the NCS in P1 is identical to the crystallographic symmetry in R32. Best, Herman *Von:*CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] *Im Auftrag von *Boaz Shaanan *Gesendet:* Mittwoch, 1. Juli 2015 12:10 *An:* CCP4BB@JISCMAIL.AC.UK mailto:CCP4BB@JISCMAIL.AC.UK *Betreff:* Re: [ccp4bb] Rfree below Rwork Just wondering about Eleanor's interesting remark: would the Rf Rw go as low as reported by Wolfram (0.22) in case of a wrong space group? Boaz /Boaz Shaanan, Ph.D. // /Dept. of Life Sciences / /Ben-Gurion University of the Negev / /Beer-Sheva 84105 / /Israel / // /E-mail: bshaa...@bgu.ac.il mailto:bshaa...@bgu.ac.il/ /Phone: 972-8-647-2220 Skype: boaz.shaanan / /Fax: 972-8-647-2992 or 972-8-646-1710 // // *From:*CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] on behalf of Eleanor Dodson [eleanor.dod...@york.ac.uk] *Sent:* Tuesday, June 30, 2015 8:55 PM *To:* CCP4BB@JISCMAIL.AC.UK mailto:CCP4BB@JISCMAIL.AC.UK *Subject:* Re: [ccp4bb] Rfree below Rwork I suppose if I was the referee for this structure and your FreeR is so close to the Rfactor I would ask you to ensure you had the right space group - is the 6 fold NCS actually 2 fold NCS with a crystallographic 3 fold.. Cases occur where R32 is indexed as C2.. Certainly if the Rfree set is assigned randomly to reflections which are symmetry equivalents then you see this phenomena of Rfree = Rfactor Eleanor On 30 June 2015 at 18:26, Gerard Bricogne g...@globalphasing.com mailto:g...@globalphasing.com wrote: Dear Wolfram, I have a perhaps optimistic view of the effect of high-order NCS on Rfree, in the sense that I don't view it as a problem. People have agonised to extreme degrees over the difficulty of choosing a free set of reflections that would produce the expected gap between Rwork and Rfree, and some of the conclusions were that you would need to hide almost half of your data in some cases! I think it is best to remember that the idea of cross-validation by Rfree is to prevent overfitting, i.e. ending up with a model that fits the amplitudes too well compared to how well it determines the phases. In the case of high-order NCS (in your case, the U/V ratio that the old papers on NCS identified as the key quantity to measure the phasing power of NCS would be less than 0.1!) the phases and the amplitudes are so tightly coupled that it is simply impossible to fit the amplitudes without delivering phases of an equally good quality. In other words there is no overfitting problem (provided you do have good and complete data) and the difference between Rfree and Rwork is simply within the bounds of the statistical spread of Rfree depending on the free set chosen. You are lucky to have 6-fold NCS, so don't let any reviewer convince you that it is a curse, and make you suffer for it :-) . With best wishes, Gerard. -- On Tue, Jun 30, 2015 at 12:58:44PM -0400, wtempel wrote: Hello, my question concerns refinement of a structure with 6-fold NCS (local automatic restraints in REFMAC) against 2.8 A data. The size of my free set is 1172 selected in thin resolution shells (SFTOOLS) and corresponding to 4.3 % of reflections. A refmac run of 10 cycles of TLS and 10 cycles of CGMAT starts out at Rfree/Rcryst 0.271/0.272. After the 10th TLS cycle I have 0.227/0.224. Yes, Rfree Rcryst. At the end of CGMAT I have 0.2072/0.2071. I understand that NCS
[ccp4bb] Post doctoral position in SAXS at EMBL Hamburg
Dear All, I would like to inform you that a postdoctoral position is available at the EMBL Hamburg Unit in the research group of Dmitri Svergun. I attach a description of the Vacancy Notice below. Deadline for application is August 15th. -- *Job description*** The European Molecular Biology Laboratory (EMBL) is one of the highest ranked scientific research organisations in the world. The Headquarters Laboratory is located in Heidelberg (Germany), with additional sites in Grenoble (France), Hamburg (Germany), Hinxton (UK) and Monterotondo (Italy). The EMBL Hamburg Outstation at the DESY campus performs research in structural biology with specific emphasis on the use of synchrotron radiation. EMBL Hamburg runs two macromolecular crystallography (MX) beamlines and a Small Angle X-ray scattering (SAXS) beamline at the high-brillance storage ring PETRA-3.The EMBL BioSAXS group of the EMBL has a strong experience in SAXS data collection, analysis and interpretation, and runs a dedicated biological SAXS beamline on Petra-3 ring providing support for over a hundred groups per year. Applications are invited for a post-doctoral position in the BioSAXS group in the frame of a collaborative project with research groups from Germany and Chech Republic. The project, supported by a grant from the German Research Foundation (DFG), deals with the SAXS studies of nanoparticles - anticancer drug carriers. The successful candidate will be involved in the collaborative work on the grant, in the operation of the beamline and in the development of new experimental approaches and biological sample environments including, in particular, anomalous and time resolved scattering on biological specimens. ** *Qualifications and Experience* Applicants should have a PhD in a relevant field (physics, engineering, structural biology) and experience with synchrotron X-ray scattering. We specifically welcome applicants with a technical background possessing a hands-on expertise in X-ray instrumentation and in biological scattering/diffraction. Good communication skills, fluent English and ability to collaborate with international teams are essential. *Application instruction* Please apply online through www.embl.org./jobs http://www.embl.org./jobs, _ _vacancy number HH_00086. Closing date : August, 15, 2015 ** *Additional Information* Further information: Dr Dmitri Svergun, email svergun(at)embl-hamburg.de Dr Clement Blanchet, email blanchet(at)embl-hamburg.de EMBL is an inclusive, equal opportunity employer offering attractive conditions and benefits appropriate to an international research organisation. The position is initially for 2 years and the appointment can be renewed, depending on circumstances at the time of the review. -- Clement Blanchet, Ph.D. EMBL Hamburg Notkestrasse 85,Bldg 25A 22607 Hamburg Germany Tel: +49 40 89902-192
Re: [ccp4bb] [phenixbb] accept MR solution even if the number of clashes is above the threshold
Dear Almu, In the development version of Phaser currently available in nightly builds of Phenix (and soon in an upcoming stable release of Phenix, as well as in CCP4), solutions that fail to pack, even though they have such high TFZ scores that they should normally have been convincing, are subjected to a pruning algorithm, where the local occupancies of residues along the chain are refined and ones that refine to low values are trimmed out. This will often sort out problems from loops, or even domains, that are in the wrong conformation. By default, potential solutions have to have a TFZ8 to be subjected to this algorithm, but this threshold can be changed. Please give this a try and tell us how it works for you! If it doesn’t, there are other possible issues, generally coming down to having the wrong spacegroup, so it might be worth double-checking the choice of spacegroup. Best wishes, Randy Read - Randy J. Read Department of Haematology, University of Cambridge Cambridge Institute for Medical ResearchTel: +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 On 2 Jul 2015, at 13:55, Almudena Ponce Salvatierra maps.fa...@gmail.com wrote: Hi everyone, I am running Phaser (from Phenix) and while checking the .log file (it is still running) I realize that it found some solutions with a TFZ score over 7, but it won't take them I guess because the number of clashes is higher than allowed (12, I guess they're not so many either). My models are poor and for so I would like to check these solutions that for now Phaser will reject. However in the input I don't want to increase the allowed number of clashes because this will then affect all the other solutions and this will take forever... So my question is: can I tell phaser somehow to write down/save the solutions for which the TFZ score will be higher than 7? Even though the clashes Thanks a lot in advance All the best, Almu -- Almudena Ponce-Salvatierra Macromolecular crystallography and Nucleic acid chemistry Max Planck Institute for Biophysical Chemistry Am Fassberg 11 37077 Göttingen Germany ___ phenixbb mailing list pheni...@phenix-online.org http://phenix-online.org/mailman/listinfo/phenixbb Unsubscribe: phenixbb-le...@phenix-online.org
Re: [ccp4bb] Problem with ViewHKL 1.08
Dear Marta, as a workaround, before starting ViewHKL change locale settings to have '.' as a decimal point. For example, from command line: LC_ALL=C viewhkl or LC_NUMERIC=C viewhkl It's a bug in ViewHKL and/or in symop_to_mat4() in libccp4. Marcin On Thu, Jul 02, 2015 at 11:43:08AM +0200, Marta Sanz Gaitero wrote: Dear all; I have reinstalled ccp4i recently and everything seems to work fine but ViewHKL. Every time I try to visualize a .mtz file the program gets opened but without loading the .mtz. Then it gets frozen and I have to kill it. In the command line the messages I get are: **SYMMETRY OPERATOR ERROR** Error in interpreting symop -X+1/2, -Y, Z+1/2 CCP4 library signal ccp4_parser:Failed to interpret symop string (Success) raised in symop_to_mat4 **SYMMETRY OPERATOR ERROR** Error in interpreting symop X+1/2, -Y+1/2, -Z CCP4 library signal ccp4_parser:Failed to interpret symop string (Success) raised in symop_to_mat4 **SYMMETRY OPERATOR ERROR** Error in interpreting symop -X, Y+1/2, -Z+1/2 CCP4 library signal ccp4_parser:Failed to interpret symop string (Success) raised in symop_to_mat4 Spacegroup_registry: ASU warning, LGhash=0x33b45b40 Spacegroup_registry: ASU fail File /media/marta/TOSHIBA/Ciencia/17_koji/20150626Xaloc/ch186/x4/b2x4/ccp4irepro/b2x4_3_0001_scaled1_refmac1.mtz is not an MTZ file My operative system is Linux Mint 17 (Qiana). Does some of you have any idea of what may be happening and how I can solve it? Thank you very much, Best regards Marta Sanz -- This e-mail and any attachments may contain confidential, copyright and or privileged material, and are for the use of the intended addressee only. If you are not the intended addressee or an authorised recipient of the addressee please notify us of receipt by returning the e-mail and do not use, copy, retain, distribute or disclose the information in or attached to the e-mail. Any opinions expressed within this e-mail are those of the individual and not necessarily of Diamond Light Source Ltd. Diamond Light Source Ltd. cannot guarantee that this e-mail or any attachments are free from viruses and we cannot accept liability for any damage which you may sustain as a result of software viruses which may be transmitted in or with the message. Diamond Light Source Limited (company no. 4375679). Registered in England and Wales with its registered office at Diamond House, Harwell Science and Innovation Campus, Didcot, Oxfordshire, OX11 0DE, United Kingdom
[ccp4bb] AW: [ccp4bb] accept MR solution even if the number of clashes is above the threshold
Dear Almu, Apparently, you have more than one search model. What worked very well for me was to superimpose these models and cut out the loops which are variable between the models. These loops may be different again in your protein and may be behind the clashes in Phaser. Without these troublesome loops, Phaser should be able to find the correct solution. You could also use all models at once, weighted by the sequence identity as explained in the Phaser manual. This might give superior results when your starting models are poor. Good luck! Herman Von: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] Im Auftrag von Almudena Ponce Salvatierra Gesendet: Donnerstag, 2. Juli 2015 14:56 An: CCP4BB@JISCMAIL.AC.UK Betreff: [ccp4bb] accept MR solution even if the number of clashes is above the threshold Hi everyone, I am running Phaser (from Phenix) and while checking the .log file (it is still running) I realize that it found some solutions with a TFZ score over 7, but it won't take them I guess because the number of clashes is higher than allowed (12, I guess they're not so many either). My models are poor and for so I would like to check these solutions that for now Phaser will reject. However in the input I don't want to increase the allowed number of clashes because this will then affect all the other solutions and this will take forever... So my question is: can I tell phaser somehow to write down/save the solutions for which the TFZ score will be higher than 7? Even though the clashes Thanks a lot in advance All the best, Almu -- Almudena Ponce-Salvatierra Macromolecular crystallography and Nucleic acid chemistry Max Planck Institute for Biophysical Chemistry Am Fassberg 11 37077 Göttingen Germany
Re: [ccp4bb] Rfree below Rwork
Hi Herman, While you're correct regarding increase in number of entities in the asu upon lowering the symmetry, you're not correct for specific case of R32. One molecule per asu in R32 equals 18 molecules per asu in P1. Regards, Vaheh Oganesyan www.medimmune.com From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of herman.schreu...@sanofi.com Sent: Wednesday, July 01, 2015 7:34 AM To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] AW: [ccp4bb] Rfree below Rwork Dear Boaz, One can equally well describe a R32 crystal with one molecule in the asymmetric unit as P1 and 6 molecules in the asymmetric unit. In this case, the NCS in P1 is identical to the crystallographic symmetry in R32. Best, Herman Von: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] Im Auftrag von Boaz Shaanan Gesendet: Mittwoch, 1. Juli 2015 12:10 An: CCP4BB@JISCMAIL.AC.UKmailto:CCP4BB@JISCMAIL.AC.UK Betreff: Re: [ccp4bb] Rfree below Rwork Just wondering about Eleanor's interesting remark: would the Rf Rw go as low as reported by Wolfram (0.22) in case of a wrong space group? Boaz Boaz Shaanan, Ph.D. Dept. of Life Sciences Ben-Gurion University of the Negev Beer-Sheva 84105 Israel E-mail: bshaa...@bgu.ac.ilmailto:bshaa...@bgu.ac.il Phone: 972-8-647-2220 Skype: boaz.shaanan Fax: 972-8-647-2992 or 972-8-646-1710 From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] on behalf of Eleanor Dodson [eleanor.dod...@york.ac.uk] Sent: Tuesday, June 30, 2015 8:55 PM To: CCP4BB@JISCMAIL.AC.UKmailto:CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] Rfree below Rwork I suppose if I was the referee for this structure and your FreeR is so close to the Rfactor I would ask you to ensure you had the right space group - is the 6 fold NCS actually 2 fold NCS with a crystallographic 3 fold.. Cases occur where R32 is indexed as C2.. Certainly if the Rfree set is assigned randomly to reflections which are symmetry equivalents then you see this phenomena of Rfree = Rfactor Eleanor On 30 June 2015 at 18:26, Gerard Bricogne g...@globalphasing.commailto:g...@globalphasing.com wrote: Dear Wolfram, I have a perhaps optimistic view of the effect of high-order NCS on Rfree, in the sense that I don't view it as a problem. People have agonised to extreme degrees over the difficulty of choosing a free set of reflections that would produce the expected gap between Rwork and Rfree, and some of the conclusions were that you would need to hide almost half of your data in some cases! I think it is best to remember that the idea of cross-validation by Rfree is to prevent overfitting, i.e. ending up with a model that fits the amplitudes too well compared to how well it determines the phases. In the case of high-order NCS (in your case, the U/V ratio that the old papers on NCS identified as the key quantity to measure the phasing power of NCS would be less than 0.1!) the phases and the amplitudes are so tightly coupled that it is simply impossible to fit the amplitudes without delivering phases of an equally good quality. In other words there is no overfitting problem (provided you do have good and complete data) and the difference between Rfree and Rwork is simply within the bounds of the statistical spread of Rfree depending on the free set chosen. You are lucky to have 6-fold NCS, so don't let any reviewer convince you that it is a curse, and make you suffer for it :-) . With best wishes, Gerard. -- On Tue, Jun 30, 2015 at 12:58:44PM -0400, wtempel wrote: Hello, my question concerns refinement of a structure with 6-fold NCS (local automatic restraints in REFMAC) against 2.8 A data. The size of my free set is 1172 selected in thin resolution shells (SFTOOLS) and corresponding to 4.3 % of reflections. A refmac run of 10 cycles of TLS and 10 cycles of CGMAT starts out at Rfree/Rcryst 0.271/0.272. After the 10th TLS cycle I have 0.227/0.224. Yes, Rfree Rcryst. At the end of CGMAT I have 0.2072/0.2071. I understand that NCS stresses the independence assumption of the free set. Am I correct in believing that Rfree *may* be smaller than Rcryst even in the absence of a major mistake? My hope is that the combined wisdom of ccp4bb followers can point out my possible mistake, suggest tests that I may perform to avoid them and, possibly, arguments in defense of a crystallographic model with Rfree Rcryst. Many thanks, Wolfram Tempel -- === * * * Gerard Bricogne g...@globalphasing.commailto:g...@globalphasing.com * * * * Global Phasing Ltd. * * Sheraton House, Castle Park Tel: +44-(0)1223-353033tel:%2B44-%280%291223-353033 * * Cambridge CB3 0AX, UK Fax:
Re: [ccp4bb] AW: [ccp4bb] Rfree below Rwork
Dirk, you're right. With rhombohedral setting there are only six copies of asymmetric units in the unit cell. So, technically, Herman was not wrong. Regards, Vaheh Oganesyan www.medimmune.com From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Dirk Kostrewa Sent: Thursday, July 02, 2015 10:03 AM To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] AW: [ccp4bb] Rfree below Rwork Hi Herman and Boaz, in the trigonal setting R32 (not in the hexagonal setting H32), the unit cell in R32 contains 6 copies. If you take the whole R32 unit cell as a P1 cell, you would have 6 copies in the asymmetric unit, as Hermann wrote. Best regards, Dirk. Am 02.07.15 um 15:52 schrieb herman.schreu...@sanofi.commailto:herman.schreu...@sanofi.com: You are right. After I sent the email to the bulletin board, I realized that in R32 there must be more then unit cells but did not send a correction. Next time, I will check the space group before sending an email. Best regards, Herman Von: Oganesyan, Vaheh [mailto:oganesy...@medimmune.com] Gesendet: Donnerstag, 2. Juli 2015 15:48 An: Schreuder, Herman RD/DE; CCP4BB@JISCMAIL.AC.UKmailto:CCP4BB@JISCMAIL.AC.UK Betreff: RE: [ccp4bb] Rfree below Rwork Hi Herman, While you're correct regarding increase in number of entities in the asu upon lowering the symmetry, you're not correct for specific case of R32. One molecule per asu in R32 equals 18 molecules per asu in P1. Regards, Vaheh Oganesyan www.medimmune.com From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of herman.schreu...@sanofi.commailto:herman.schreu...@sanofi.com Sent: Wednesday, July 01, 2015 7:34 AM To: CCP4BB@JISCMAIL.AC.UKmailto:CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] AW: [ccp4bb] Rfree below Rwork Dear Boaz, One can equally well describe a R32 crystal with one molecule in the asymmetric unit as P1 and 6 molecules in the asymmetric unit. In this case, the NCS in P1 is identical to the crystallographic symmetry in R32. Best, Herman Von: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] Im Auftrag von Boaz Shaanan Gesendet: Mittwoch, 1. Juli 2015 12:10 An: CCP4BB@JISCMAIL.AC.UKmailto:CCP4BB@JISCMAIL.AC.UK Betreff: Re: [ccp4bb] Rfree below Rwork Just wondering about Eleanor's interesting remark: would the Rf Rw go as low as reported by Wolfram (0.22) in case of a wrong space group? Boaz Boaz Shaanan, Ph.D. Dept. of Life Sciences Ben-Gurion University of the Negev Beer-Sheva 84105 Israel E-mail: bshaa...@bgu.ac.ilmailto:bshaa...@bgu.ac.il Phone: 972-8-647-2220 Skype: boaz.shaanan Fax: 972-8-647-2992 or 972-8-646-1710 From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UKmailto:CCP4BB@JISCMAIL.AC.UK] on behalf of Eleanor Dodson [eleanor.dod...@york.ac.ukmailto:eleanor.dod...@york.ac.uk] Sent: Tuesday, June 30, 2015 8:55 PM To: CCP4BB@JISCMAIL.AC.UKmailto:CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] Rfree below Rwork I suppose if I was the referee for this structure and your FreeR is so close to the Rfactor I would ask you to ensure you had the right space group - is the 6 fold NCS actually 2 fold NCS with a crystallographic 3 fold.. Cases occur where R32 is indexed as C2.. Certainly if the Rfree set is assigned randomly to reflections which are symmetry equivalents then you see this phenomena of Rfree = Rfactor Eleanor On 30 June 2015 at 18:26, Gerard Bricogne g...@globalphasing.commailto:g...@globalphasing.com wrote: Dear Wolfram, I have a perhaps optimistic view of the effect of high-order NCS on Rfree, in the sense that I don't view it as a problem. People have agonised to extreme degrees over the difficulty of choosing a free set of reflections that would produce the expected gap between Rwork and Rfree, and some of the conclusions were that you would need to hide almost half of your data in some cases! I think it is best to remember that the idea of cross-validation by Rfree is to prevent overfitting, i.e. ending up with a model that fits the amplitudes too well compared to how well it determines the phases. In the case of high-order NCS (in your case, the U/V ratio that the old papers on NCS identified as the key quantity to measure the phasing power of NCS would be less than 0.1!) the phases and the amplitudes are so tightly coupled that it is simply impossible to fit the amplitudes without delivering phases of an equally good quality. In other words there is no overfitting problem (provided you do have good and complete data) and the difference between Rfree and Rwork is simply within the bounds of the statistical spread of Rfree depending on the free set chosen. You are lucky to have 6-fold NCS, so don't let any reviewer convince you that it is a curse, and make you suffer for it :-) . With best wishes, Gerard. -- On Tue, Jun 30, 2015 at 12:58:44PM -0400, wtempel wrote: Hello, my question concerns refinement of a structure with 6-fold NCS (local automatic restraints in REFMAC)
Re: [ccp4bb] paired refinement
On Thu, 2 Jul 2015 00:15:58 +0100, Eric Karg harvard...@yahoo.com wrote: Hi all, I have a dataset processed in XDS to 2.3 A (based on CC1/2). I'm trying to do paired refinement to determine the optimal resolution cutoff. Here is what I get at different resolutions set in Phenix: Final Rfree/Rwork: 2.7— 0.2498/0.2027 2.6— 0.2519/0.2009 2.5— 0.2567/0.2025 2.4 — 0.2481/0.2042 2.3 — 0.2493/0.2075 The geometry of all output structures are similar. 1. What is the high resolution cutoff based on these data? I know that Rfree/Rwork at different resolution should not be compared, but is there a simple way to do the test as described in the KD 2012 Science paper using Phenix GUI? not, as far as I know, in a single step. Rather, you need to find out Rwork/Rfree of the 2.3A model, against the 2.4A data: if Rfree is better than 0.2481 (and Rfree-Rwork 0.2481-0.2042), that would tell you that the reflections between 2.4 and 2.3 A improve the model. For consistency, I would then expect that a similar calculation involving the 2.3A model and the 2.5A data _also_ shows that the 2.3A model is superior than the 2.5A model. That would be two steps, but the second is of course not mandatory. If, on the other hand, Rfree of the 2.3A model is worse than 0.2481 against the 2.4A data, you should disregard 2.3A as a potential high-resolution cutoff. In that case, you would continue along the same lines, by using the 2.4A model, and investigating its Rwork/Rfree against the 2.5A data. 2. For refining a structure at a lower resolution (lower than the initial dataset), do I simply set the resolution limit in the refinement or I need to reprocess the data starting from the images? Do I need to do anything with Rfree flags? Based on the discussions on this forum I know I should deposit the highest resolution dataset but my question is about the mtz file which will be used for refinement. All you need at a lower resolution is Rwork/Rfree (of the higher-resolution model) _without_ positional or B-factor refinement. It is easiest to run (on the command line; don't know how to do it in the GUI) phenix.model_vs_data high_resolution=2.4 project.pdb project.mtz and finding the values of r_work_cutoff and r_free_cutoff in the output. These values are close (within 0.2%) to those you would obtain from a run of phenix.refine with xray_data.high_resolution=2.4 main.number_of_macro_cycles=0 . HTH, Kay Thank you very much for your help!
Re: [ccp4bb] ssDNA
1) Commercially made oligonucleotides 2) M13 rolling circle replication (how people used to make ssDNA for sanger sequencing). ——— Dr Antony W Oliver Senior Research Fellow CR-UK DNA Repair Enzymes Group Genome Damage and Stability Centre Science Park Road University of Sussex Falmer, Brighton, BN1 9RQ e: antony.oli...@sussex.ac.ukmailto:antony.oli...@sussex.ac.uk t(office): +44 (0)1273 678349 t(lab): +44 (0)1273 677512 http://www.sussex.ac.uk/lifesci/oliverlab http://tinyurl.com/aw-oliver [cid:EF31410E-E250-44AB-83E7-4BCEAF6FDAD0@gdsc.susx.ac.uk] On 2 Jul 2015, at 16:40, Reza Khayat rkha...@ccny.cuny.edumailto:rkha...@ccny.cuny.edu wrote: Hi, Sorry for the non-crystallography question, does anyone know how to produce milligram quantities of single stranded DNA? Thanks. Best wishes, Reza Reza Khayat, PhD Assistant Professor Department of Chemistry City College of New York 85 Saint Nicholas Terrace, CDI 12318 New York, NY 10031 http://www.khayatlab.org/ 212-650-6070
[ccp4bb] ssDNA
Hi, Sorry for the non-crystallography question, does anyone know how to produce milligram quantities of single stranded DNA? Thanks. Best wishes, Reza Reza Khayat, PhD Assistant Professor Department of Chemistry City College of New York 85 Saint Nicholas Terrace, CDI 12318 New York, NY 10031 http://www.khayatlab.org/ 212-650-6070
Re: [ccp4bb] paired refinement
I think your idea was discussed herein before, but that the consensus was that CC1/2 does not change much when you scramble and re-calculate. What one needs is a good way to determine at what point in the CC1/2 curve the data stops being useful in obtaining the best possible model, and it seems that this point may depend on individual cases. In a way, there is still some information in reflections with I/sigma approaching one, since even those measurements limit the possible values of I if only slightly, i.e., the true value of I can't be greater than 1000 or something. I think that if crystallography still exists in 20 years, we will be refining directly against pixel/voxel/roxel(?) intensities. JPK -Original Message- From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Takanori Nakane Sent: Thursday, July 02, 2015 8:53 PM To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] paired refinement Hi all, I have read recent SERCA paper on IUCrJ and found their discussion interesting. Structural studies of P-type ATPase–ligand complexes using an X-ray free-electron laser by Maike Bublitz et al. http://journals.iucr.org/m/issues/2015/04/00/jt5009/index.html In addition to CC1/2 and paired refinement, they scrambled indices of reflections and showed Rfree indeed went up. If we repeat permutations many times and calculate CC1/2 every time, we can obtain empirical distribution of CC1/2 under null hypothesis and thus calculate p-value. This is called permutation test in statistics. I am wondering how this p-value compares with p-values calculated by Student's t-test. Since t-test does not take the effect of multiplicity into the account, I think the permutation test gives more accurate value. (That is, CC1/2 calculated from a dataset with multiplicity 30 should be more accurate and reliable than that calculated from a dataset with multiplicity 2, but this difference is ignored in t-test.) Did anyone try this? Best regards, Takanori Nakane On 2015年07月03日 04:41, Tim Gruene wrote: Hi Robbie, I have been wondering how much information would be present in a weighted CC1/2 with weights from the ML refinement program. As I understand the concept behind paired refinement, one can use much higher resolution data in refinement than you would expect from the (classical) scaling statistics, because refinement programs down-weight the outliers. Maybe a weighted CC1/2 would be a convenient short cut the more time consuming paired refinemement? How easily could the calculation of a weighted CC1/2 be implemented in refmac? Refmac would have to merge the data internally, but that's just a simple formula to implement. Best, Tim On 07/02/2015 08:28 PM, Robbie Joosten wrote: Hi Jacob, You need unmerged data to calculate cc1/2. That's not the sort of data you get from the PDB. But anyway, we have a fairly simple automated test that we can use on a case-by-case basis. I would argue that that is nicer than a empirical cut-off that may or may not be correct for you case. Cheers, Robbie Sent with my Windows Phone Van: Keller, Jacobmailto:kell...@janelia.hhmi.org Verzonden: 2-7-2015 20:12 Aan: CCP4BB@JISCMAIL.AC.UKmailto:CCP4BB@JISCMAIL.AC.UK Onderwerp: Re: [ccp4bb] paired refinement Well, in that case, one could simply look at the plot of CC1/2 versus resolution and see the step up to one, conclude something was off. I wonder whether PDB REDO was able to get some empirically-determined values for CC1/2 cutoffs by comparing paired refinement versus CC1/2 versus other parameters? JPK -Original Message- From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Dale Tronrud Sent: Thursday, July 02, 2015 1:46 PM To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] paired refinement While I was puzzling over an entry in the PDB some years ago (since obsoleted) I noticed that all the high resolution amplitudes were equal to 11.0! This was before CC1/2 but for this structure it would have been equal to one, and yet the outer data were useless. A practical test like paired refinement can't be fooled in this way. Dale Tronrud On 7/2/2015 10:25 AM, Edward A. Berry wrote: My take on this- No one has been willing to specify a cutoff (and probably there is no rigorous way to mathematically define the cutoff) and say If CC* (or CCfree or whatever) is below X then it will not improve your structure, if above X then it will. Probably depends among other things on how strong the lower resolution data is, how good the structure is without the added data. On the other hand in paired refinement, if adding the data improves the structure as measured by Rfree in a zone excluding the added data, then it is hard to deny that that data are worth including. eab On 07/02/2015 12:52 PM, Keller, Jacob wrote: Wasn’t all of this put to bed through the implementation
Re: [ccp4bb] paired refinement
Hi all, I have read recent SERCA paper on IUCrJ and found their discussion interesting. Structural studies of P-type ATPase–ligand complexes using an X-ray free-electron laser by Maike Bublitz et al. http://journals.iucr.org/m/issues/2015/04/00/jt5009/index.html In addition to CC1/2 and paired refinement, they scrambled indices of reflections and showed Rfree indeed went up. If we repeat permutations many times and calculate CC1/2 every time, we can obtain empirical distribution of CC1/2 under null hypothesis and thus calculate p-value. This is called permutation test in statistics. I am wondering how this p-value compares with p-values calculated by Student's t-test. Since t-test does not take the effect of multiplicity into the account, I think the permutation test gives more accurate value. (That is, CC1/2 calculated from a dataset with multiplicity 30 should be more accurate and reliable than that calculated from a dataset with multiplicity 2, but this difference is ignored in t-test.) Did anyone try this? Best regards, Takanori Nakane On 2015年07月03日 04:41, Tim Gruene wrote: Hi Robbie, I have been wondering how much information would be present in a weighted CC1/2 with weights from the ML refinement program. As I understand the concept behind paired refinement, one can use much higher resolution data in refinement than you would expect from the (classical) scaling statistics, because refinement programs down-weight the outliers. Maybe a weighted CC1/2 would be a convenient short cut the more time consuming paired refinemement? How easily could the calculation of a weighted CC1/2 be implemented in refmac? Refmac would have to merge the data internally, but that's just a simple formula to implement. Best, Tim On 07/02/2015 08:28 PM, Robbie Joosten wrote: Hi Jacob, You need unmerged data to calculate cc1/2. That's not the sort of data you get from the PDB. But anyway, we have a fairly simple automated test that we can use on a case-by-case basis. I would argue that that is nicer than a empirical cut-off that may or may not be correct for you case. Cheers, Robbie Sent with my Windows Phone Van: Keller, Jacobmailto:kell...@janelia.hhmi.org Verzonden: 2-7-2015 20:12 Aan: CCP4BB@JISCMAIL.AC.UKmailto:CCP4BB@JISCMAIL.AC.UK Onderwerp: Re: [ccp4bb] paired refinement Well, in that case, one could simply look at the plot of CC1/2 versus resolution and see the step up to one, conclude something was off. I wonder whether PDB REDO was able to get some empirically-determined values for CC1/2 cutoffs by comparing paired refinement versus CC1/2 versus other parameters? JPK -Original Message- From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Dale Tronrud Sent: Thursday, July 02, 2015 1:46 PM To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] paired refinement While I was puzzling over an entry in the PDB some years ago (since obsoleted) I noticed that all the high resolution amplitudes were equal to 11.0! This was before CC1/2 but for this structure it would have been equal to one, and yet the outer data were useless. A practical test like paired refinement can't be fooled in this way. Dale Tronrud On 7/2/2015 10:25 AM, Edward A. Berry wrote: My take on this- No one has been willing to specify a cutoff (and probably there is no rigorous way to mathematically define the cutoff) and say If CC* (or CCfree or whatever) is below X then it will not improve your structure, if above X then it will. Probably depends among other things on how strong the lower resolution data is, how good the structure is without the added data. On the other hand in paired refinement, if adding the data improves the structure as measured by Rfree in a zone excluding the added data, then it is hard to deny that that data are worth including. eab On 07/02/2015 12:52 PM, Keller, Jacob wrote: Wasn’t all of this put to bed through the implementation of CC measures? JPK *From:*CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] *On Behalf Of *Robbie Joosten *Sent:* Thursday, July 02, 2015 12:46 PM *To:* CCP4BB@JISCMAIL.AC.UK *Subject:* Re: [ccp4bb] paired refinement But it is not the R-free of the shell here. In paired refinement you take the R-free of the reflections outside the shell. Cheers, Robbie Sent with my Windows Phone - - - - - - - -
Re: [ccp4bb] Coot and Pymol through SSH by Xming/PUTTY on a windows client?
Thanks for everybody's suggestions! I finally fixed coot following Zhijie's suggestion, where I set one window mode in Xlaunch. For pymol, I just gave up... I am not sure whether the NVIDIA graphic card is making things more complicated... Best, Chen On Thu, Jul 2, 2015 at 1:29 AM, Zhijie Li zhijie...@utoronto.ca wrote: Hi Chen, I followed instructions on this page and it seems to be working: http://www.geo.mtu.edu/geoschem/docs/putty_install.html https://urldefense.proofpoint.com/v2/url?u=http-3A__www.geo.mtu.edu_geoschem_docs_putty-5Finstall.htmld=AwMFaQc=-dg2m7zWuuDZ0MUcV7Sdqwr=uV9bK9zAIvRZlk7q6-YllAm=xfQGepyCVehabnXteqeg61Xem6P7gGu-1ba56BOC5wcs=cgZAn_6jKDjQ-qmii2I1r5svq0nA403oEVm7Mite95Ie= One thing I think is worth mentioning is that in Putty-connection-SSH-X11, I put localhost:0.0 instead of 10.0, as for putty itself, it is requesting from Xming for the use of display 0.0. For COOT, it seems that I have to set Xlaunch in “one window” mode or simply run Xming.exe. But still there is some issue with refreshing the screen when choosing menu items. My systems are CentOS 6.6 and Windows 7/Putty0.60/Xming6.9.0.31. Zhijie *From:* Chen Zhao c.z...@yale.edu *Sent:* Wednesday, July 01, 2015 7:05 PM *To:* CCP4BB@JISCMAIL.AC.UK *Subject:* Re: [ccp4bb] Coot and Pymol through SSH by Xming/PUTTY on a windows client? Thank you Dale, but when I added localhost:10.0 to X display location, the problem still exist, just without the phrase localhost:10.0 in the warning. My X11 forwarding is enabled all the time and all other GUIs work just fine. And thank you for your clarification on the concept of server and client in the X11 word. It makes a lot of sense and I just didn't give it a second thought! Best, Chen On Wed, Jul 1, 2015 at 6:47 PM, Dale Tronrud de...@daletronrud.com wrote: -BEGIN PGP SIGNED MESSAGE- Hash: SHA1 Both the ssh client and server must be set up with X11Forwarding yes. The message sounds like your local computer is not set up to accept X11 tunneling. (By the way, in the X11 world the remote system is the client and your local system the server.) Dale Tronrud On 7/1/2015 3:40 PM, Chen Zhao wrote: Hi all, Sorry to bother you, but I am trying to fix a long-standing problem that I cannot run Coot and Pymol through Xming/PUTTY by SSH connection on a windows client. The error messages are pretty similar for both: Coot: PuTTY X11 proxy: unable to connect to forwarded X server: Network error: Connection refused (coot-real:23113): Gtk-WARNING **: cannot open display: localhost:10.0 Pymol: PuTTY X11 proxy: unable to connect to forwarded X server: Network error: Connection refused freeglut (pymol): failed to open display 'localhost:10.0' Does anybody have some ideas? Thank you so much, Chen -BEGIN PGP SIGNATURE- Version: GnuPG v2.0.22 (MingW32) iEYEARECAAYFAlWUbgYACgkQU5C0gGfAG10hVgCeLmuE4pHFrapu9biY9nHO/Bpi 5O8An17UN+hgpr7/6A+mny+XOBfJV/T5 =iTfz -END PGP SIGNATURE-
Re: [ccp4bb] paired refinement
Another criterion for cutoff, also requiring the structure to be solved, is the agreement between data and structure, e.g. Rfree or CCfree. I think it is very unlikely that you could get Rfree =.2493 in a shell which contains only noise. So I would suggest doing paired refinement to 2.2 and 2.1 A (if the data is available). On 07/01/2015 07:15 PM, Eric Karg wrote: Hi all, I have a dataset processed in XDS to 2.3 A (based on CC1/2). I'm trying to do paired refinement to determine the optimal resolution cutoff. Here is what I get at different resolutions set in Phenix: Final Rfree/Rwork: 2.7— 0.2498/0.2027 2.6— 0.2519/0.2009 2.5— 0.2567/0.2025 2.4 — 0.2481/0.2042 2.3 — 0.2493/0.2075 The geometry of all output structures are similar. 1. What is the high resolution cutoff based on these data? I know that Rfree/Rwork at different resolution should not be compared, but is there a simple way to do the test as described in the KD 2012 Science paper using Phenix GUI? 2. For refining a structure at a lower resolution (lower than the initial dataset), do I simply set the resolution limit in the refinement or I need to reprocess the data starting from the images? Do I need to do anything with Rfree flags? Based on the discussions on this forum I know I should deposit the highest resolution dataset but my question is about the mtz file which will be used for refinement. Thank you very much for your help!
Re: [ccp4bb] CCP4 setup (LIBCHECK)
On 02/07/15 18:01, Pankaj Chauhan wrote: I have installed ccp4 and phenix on ubuntu 14.04 LTS. When I open coot while using phenix or ccp4 and try to load a molecule from a Search Monomer library, it is unable to load the molecule and this pop up You need to setup CCP4 (Specifically LIBCHECK) first. Old Coots used to say this. I don't believe the current one does (unless the ligand doesn't have coordinates in the dictionary). When asking coot questions is generally a good idea to make a note of the version and from where you downloaded it. Regards, Paul.
Re: [ccp4bb] paired refinement
Well, in that case, one could simply look at the plot of CC1/2 versus resolution and see the step up to one, conclude something was off. I wonder whether PDB REDO was able to get some empirically-determined values for CC1/2 cutoffs by comparing paired refinement versus CC1/2 versus other parameters? JPK -Original Message- From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Dale Tronrud Sent: Thursday, July 02, 2015 1:46 PM To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] paired refinement While I was puzzling over an entry in the PDB some years ago (since obsoleted) I noticed that all the high resolution amplitudes were equal to 11.0! This was before CC1/2 but for this structure it would have been equal to one, and yet the outer data were useless. A practical test like paired refinement can't be fooled in this way. Dale Tronrud On 7/2/2015 10:25 AM, Edward A. Berry wrote: My take on this- No one has been willing to specify a cutoff (and probably there is no rigorous way to mathematically define the cutoff) and say If CC* (or CCfree or whatever) is below X then it will not improve your structure, if above X then it will. Probably depends among other things on how strong the lower resolution data is, how good the structure is without the added data. On the other hand in paired refinement, if adding the data improves the structure as measured by Rfree in a zone excluding the added data, then it is hard to deny that that data are worth including. eab On 07/02/2015 12:52 PM, Keller, Jacob wrote: Wasn’t all of this put to bed through the implementation of CC measures? JPK *From:*CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] *On Behalf Of *Robbie Joosten *Sent:* Thursday, July 02, 2015 12:46 PM *To:* CCP4BB@JISCMAIL.AC.UK *Subject:* Re: [ccp4bb] paired refinement But it is not the R-free of the shell here. In paired refinement you take the R-free of the reflections outside the shell. Cheers, Robbie Sent with my Windows Phone - - - - - - - - - - - - - - - --- *Van: *Edward A. Berry mailto:ber...@upstate.edu *Verzonden: *2-7-2015 18:43 *Aan: *CCP4BB@JISCMAIL.AC.UK mailto:CCP4BB@JISCMAIL.AC.UK *Onderwerp: *Re: [ccp4bb] paired refinement Another criterion for cutoff, also requiring the structure to be solved, is the agreement between data and structure, e.g. Rfree or CCfree. I think it is very unlikely that you could get Rfree =.2493 in a shell which contains only noise. So I would suggest doing paired refinement to 2.2 and 2.1 A (if the data is available). On 07/01/2015 07:15 PM, Eric Karg wrote: Hi all, I have a dataset processed in XDS to 2.3 A (based on CC1/2). I'm trying to do paired refinement to determine the optimal resolution cutoff. Here is what I get at different resolutions set in Phenix: Final Rfree/Rwork: 2.7— 0.2498/0.2027 2.6— 0.2519/0.2009 2.5— 0.2567/0.2025 2.4 — 0.2481/0.2042 2.3 — 0.2493/0.2075 The geometry of all output structures are similar. 1. What is the high resolution cutoff based on these data? I know that Rfree/Rwork at different resolution should not be compared, but is there a simple way to do the test as described in the KD 2012 Science paper using Phenix GUI? 2. For refining a structure at a lower resolution (lower than the initial dataset), do I simply set the resolution limit in the refinement or I need to reprocess the data starting from the images? Do I need to do anything with Rfree flags? Based on the discussions on this forum I know I should deposit the highest resolution dataset but my question is about the mtz file which will be used for refinement. Thank you very much for your help!
Re: [ccp4bb] paired refinement
While I was puzzling over an entry in the PDB some years ago (since obsoleted) I noticed that all the high resolution amplitudes were equal to 11.0! This was before CC1/2 but for this structure it would have been equal to one, and yet the outer data were useless. A practical test like paired refinement can't be fooled in this way. Dale Tronrud On 7/2/2015 10:25 AM, Edward A. Berry wrote: My take on this- No one has been willing to specify a cutoff (and probably there is no rigorous way to mathematically define the cutoff) and say If CC* (or CCfree or whatever) is below X then it will not improve your structure, if above X then it will. Probably depends among other things on how strong the lower resolution data is, how good the structure is without the added data. On the other hand in paired refinement, if adding the data improves the structure as measured by Rfree in a zone excluding the added data, then it is hard to deny that that data are worth including. eab On 07/02/2015 12:52 PM, Keller, Jacob wrote: Wasn’t all of this put to bed through the implementation of CC measures? JPK *From:*CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] *On Behalf Of *Robbie Joosten *Sent:* Thursday, July 02, 2015 12:46 PM *To:* CCP4BB@JISCMAIL.AC.UK *Subject:* Re: [ccp4bb] paired refinement But it is not the R-free of the shell here. In paired refinement you take the R-free of the reflections outside the shell. Cheers, Robbie Sent with my Windows Phone --- --- *Van: *Edward A. Berry mailto:ber...@upstate.edu *Verzonden: *2-7-2015 18:43 *Aan: *CCP4BB@JISCMAIL.AC.UK mailto:CCP4BB@JISCMAIL.AC.UK *Onderwerp: *Re: [ccp4bb] paired refinement Another criterion for cutoff, also requiring the structure to be solved, is the agreement between data and structure, e.g. Rfree or CCfree. I think it is very unlikely that you could get Rfree =.2493 in a shell which contains only noise. So I would suggest doing paired refinement to 2.2 and 2.1 A (if the data is available). On 07/01/2015 07:15 PM, Eric Karg wrote: Hi all, I have a dataset processed in XDS to 2.3 A (based on CC1/2). I'm trying to do paired refinement to determine the optimal resolution cutoff. Here is what I get at different resolutions set in Phenix: Final Rfree/Rwork: 2.7— 0.2498/0.2027 2.6— 0.2519/0.2009 2.5— 0.2567/0.2025 2.4 — 0.2481/0.2042 2.3 — 0.2493/0.2075 The geometry of all output structures are similar. 1. What is the high resolution cutoff based on these data? I know that Rfree/Rwork at different resolution should not be compared, but is there a simple way to do the test as described in the KD 2012 Science paper using Phenix GUI? 2. For refining a structure at a lower resolution (lower than the initial dataset), do I simply set the resolution limit in the refinement or I need to reprocess the data starting from the images? Do I need to do anything with Rfree flags? Based on the discussions on this forum I know I should deposit the highest resolution dataset but my question is about the mtz file which will be used for refinement. Thank you very much for your help!
[ccp4bb] CCP4 setup (LIBCHECK)
Hi, I have installed ccp4 and phenix on ubuntu 14.04 LTS. When I open coot while using phenix or ccp4 and try to load a molecule from a Search Monomer library, it is unable to load the molecule and this pop up You need to setup CCP4 (Specifically LIBCHECK) first. I will appreciate if someone can guide through this. Thanks Pankaj Kumar Gyanu Lemichhane lab http://webhost.nts.jhu.edu/gl/ Centre for Tuberculosis Research Lab Department of Infectious Disease Johns Hopkins University School of Medicine 725 N. Wolfe St. Baltimore, MD 21205-2185 Lab phone: (410) 955-3967 pkuma...@jhmi.edu pkuma...@commsmail.johnshopkins.edu http://pankajimtech.webs.com
Re: [ccp4bb] paired refinement
My take on this- No one has been willing to specify a cutoff (and probably there is no rigorous way to mathematically define the cutoff) and say If CC* (or CCfree or whatever) is below X then it will not improve your structure, if above X then it will. Probably depends among other things on how strong the lower resolution data is, how good the structure is without the added data. On the other hand in paired refinement, if adding the data improves the structure as measured by Rfree in a zone excluding the added data, then it is hard to deny that that data are worth including. eab On 07/02/2015 12:52 PM, Keller, Jacob wrote: Wasn’t all of this put to bed through the implementation of CC measures? JPK *From:*CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] *On Behalf Of *Robbie Joosten *Sent:* Thursday, July 02, 2015 12:46 PM *To:* CCP4BB@JISCMAIL.AC.UK *Subject:* Re: [ccp4bb] paired refinement But it is not the R-free of the shell here. In paired refinement you take the R-free of the reflections outside the shell. Cheers, Robbie Sent with my Windows Phone -- *Van: *Edward A. Berry mailto:ber...@upstate.edu *Verzonden: *2-7-2015 18:43 *Aan: *CCP4BB@JISCMAIL.AC.UK mailto:CCP4BB@JISCMAIL.AC.UK *Onderwerp: *Re: [ccp4bb] paired refinement Another criterion for cutoff, also requiring the structure to be solved, is the agreement between data and structure, e.g. Rfree or CCfree. I think it is very unlikely that you could get Rfree =.2493 in a shell which contains only noise. So I would suggest doing paired refinement to 2.2 and 2.1 A (if the data is available). On 07/01/2015 07:15 PM, Eric Karg wrote: Hi all, I have a dataset processed in XDS to 2.3 A (based on CC1/2). I'm trying to do paired refinement to determine the optimal resolution cutoff. Here is what I get at different resolutions set in Phenix: Final Rfree/Rwork: 2.7— 0.2498/0.2027 2.6— 0.2519/0.2009 2.5— 0.2567/0.2025 2.4 — 0.2481/0.2042 2.3 — 0.2493/0.2075 The geometry of all output structures are similar. 1. What is the high resolution cutoff based on these data? I know that Rfree/Rwork at different resolution should not be compared, but is there a simple way to do the test as described in the KD 2012 Science paper using Phenix GUI? 2. For refining a structure at a lower resolution (lower than the initial dataset), do I simply set the resolution limit in the refinement or I need to reprocess the data starting from the images? Do I need to do anything with Rfree flags? Based on the discussions on this forum I know I should deposit the highest resolution dataset but my question is about the mtz file which will be used for refinement. Thank you very much for your help!
Re: [ccp4bb] paired refinement
But it is not the R-free of the shell here. In paired refinement you take the R-free of the reflections outside the shell. Cheers, Robbie Sent with my Windows Phone Van: Edward A. Berrymailto:ber...@upstate.edu Verzonden: 2-7-2015 18:43 Aan: CCP4BB@JISCMAIL.AC.UKmailto:CCP4BB@JISCMAIL.AC.UK Onderwerp: Re: [ccp4bb] paired refinement Another criterion for cutoff, also requiring the structure to be solved, is the agreement between data and structure, e.g. Rfree or CCfree. I think it is very unlikely that you could get Rfree =.2493 in a shell which contains only noise. So I would suggest doing paired refinement to 2.2 and 2.1 A (if the data is available). On 07/01/2015 07:15 PM, Eric Karg wrote: Hi all, I have a dataset processed in XDS to 2.3 A (based on CC1/2). I'm trying to do paired refinement to determine the optimal resolution cutoff. Here is what I get at different resolutions set in Phenix: Final Rfree/Rwork: 2.7— 0.2498/0.2027 2.6— 0.2519/0.2009 2.5— 0.2567/0.2025 2.4 — 0.2481/0.2042 2.3 — 0.2493/0.2075 The geometry of all output structures are similar. 1. What is the high resolution cutoff based on these data? I know that Rfree/Rwork at different resolution should not be compared, but is there a simple way to do the test as described in the KD 2012 Science paper using Phenix GUI? 2. For refining a structure at a lower resolution (lower than the initial dataset), do I simply set the resolution limit in the refinement or I need to reprocess the data starting from the images? Do I need to do anything with Rfree flags? Based on the discussions on this forum I know I should deposit the highest resolution dataset but my question is about the mtz file which will be used for refinement. Thank you very much for your help!
Re: [ccp4bb] paired refinement
Yes, my stupid mistake. Please delete/disregard! On 07/02/2015 12:46 PM, Robbie Joosten wrote: But it is not the R-free of the shell here. In paired refinement you take the R-free of the reflections outside the shell. Cheers, Robbie Sent with my Windows Phone -- Van: Edward A. Berry mailto:ber...@upstate.edu Verzonden: 2-7-2015 18:43 Aan: CCP4BB@JISCMAIL.AC.UK mailto:CCP4BB@JISCMAIL.AC.UK Onderwerp: Re: [ccp4bb] paired refinement Another criterion for cutoff, also requiring the structure to be solved, is the agreement between data and structure, e.g. Rfree or CCfree. I think it is very unlikely that you could get Rfree =.2493 in a shell which contains only noise. So I would suggest doing paired refinement to 2.2 and 2.1 A (if the data is available). On 07/01/2015 07:15 PM, Eric Karg wrote: Hi all, I have a dataset processed in XDS to 2.3 A (based on CC1/2). I'm trying to do paired refinement to determine the optimal resolution cutoff. Here is what I get at different resolutions set in Phenix: Final Rfree/Rwork: 2.7— 0.2498/0.2027 2.6— 0.2519/0.2009 2.5— 0.2567/0.2025 2.4 — 0.2481/0.2042 2.3 — 0.2493/0.2075 The geometry of all output structures are similar. 1. What is the high resolution cutoff based on these data? I know that Rfree/Rwork at different resolution should not be compared, but is there a simple way to do the test as described in the KD 2012 Science paper using Phenix GUI? 2. For refining a structure at a lower resolution (lower than the initial dataset), do I simply set the resolution limit in the refinement or I need to reprocess the data starting from the images? Do I need to do anything with Rfree flags? Based on the discussions on this forum I know I should deposit the highest resolution dataset but my question is about the mtz file which will be used for refinement. Thank you very much for your help!
Re: [ccp4bb] paired refinement
Wasn’t all of this put to bed through the implementation of CC measures? JPK From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Robbie Joosten Sent: Thursday, July 02, 2015 12:46 PM To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] paired refinement But it is not the R-free of the shell here. In paired refinement you take the R-free of the reflections outside the shell. Cheers, Robbie Sent with my Windows Phone Van: Edward A. Berrymailto:ber...@upstate.edu Verzonden: 2-7-2015 18:43 Aan: CCP4BB@JISCMAIL.AC.UKmailto:CCP4BB@JISCMAIL.AC.UK Onderwerp: Re: [ccp4bb] paired refinement Another criterion for cutoff, also requiring the structure to be solved, is the agreement between data and structure, e.g. Rfree or CCfree. I think it is very unlikely that you could get Rfree =.2493 in a shell which contains only noise. So I would suggest doing paired refinement to 2.2 and 2.1 A (if the data is available). On 07/01/2015 07:15 PM, Eric Karg wrote: Hi all, I have a dataset processed in XDS to 2.3 A (based on CC1/2). I'm trying to do paired refinement to determine the optimal resolution cutoff. Here is what I get at different resolutions set in Phenix: Final Rfree/Rwork: 2.7— 0.2498/0.2027 2.6— 0.2519/0.2009 2.5— 0.2567/0.2025 2.4 — 0.2481/0.2042 2.3 — 0.2493/0.2075 The geometry of all output structures are similar. 1. What is the high resolution cutoff based on these data? I know that Rfree/Rwork at different resolution should not be compared, but is there a simple way to do the test as described in the KD 2012 Science paper using Phenix GUI? 2. For refining a structure at a lower resolution (lower than the initial dataset), do I simply set the resolution limit in the refinement or I need to reprocess the data starting from the images? Do I need to do anything with Rfree flags? Based on the discussions on this forum I know I should deposit the highest resolution dataset but my question is about the mtz file which will be used for refinement. Thank you very much for your help!
Re: [ccp4bb] CCP4 setup (LIBCHECK)
Dear Paul, The software is coot-Linux-x86_64-ubuntu-14.04. Have I made any mistake in installing or executing any file of coot during installation (there is a file, LIBEXEC). thanks Pankaj On Thu, Jul 2, 2015 at 1:41 PM, Paul Emsley pems...@mrc-lmb.cam.ac.uk wrote: On 02/07/15 18:01, Pankaj Chauhan wrote: I have installed ccp4 and phenix on ubuntu 14.04 LTS. When I open coot while using phenix or ccp4 and try to load a molecule from a Search Monomer library, it is unable to load the molecule and this pop up You need to setup CCP4 (Specifically LIBCHECK) first. Old Coots used to say this. I don't believe the current one does (unless the ligand doesn't have coordinates in the dictionary). When asking coot questions is generally a good idea to make a note of the version and from where you downloaded it. Regards, Paul.
Re: [ccp4bb] Heavy-atom derivatives
Dear Giulliana Rangel, if your protein is made recombinantly you could try a SeMet prep. The fixed atoms solve many issues you could have with soaking. Best, Tim On 06/29/2015 02:47 PM, Giulliana Rangel wrote: Dear all, I'm looking for a method to solve the phase problem. Thus, I would like some help about heavy-atom derivatives. What is the best heavy-atom to make soaking? I tried Iodine (because my protein have a lot of tyrosines), but no results... I read some papers about quick-soak, its a good method? Using platine, Hg, Br... Any suggestions? Thank you so much. Best wishes, -- -- Dr Tim Gruene Institut fuer anorganische Chemie Tammannstr. 4 D-37077 Goettingen phone: +49 (0)551 39 22149 GPG Key ID = A46BEE1A signature.asc Description: OpenPGP digital signature
Re: [ccp4bb] paired refinement
Hi Jacob, An automated test for finding a resolution cut-off: Paired refinement. I may be somewhat biased here, but I think it is fairly conveniently implement in PDB_REDO ;) Cheers, Robbie Sent with my Windows Phone Van: Keller, Jacobmailto:kell...@janelia.hhmi.org Verzonden: 2-7-2015 20:40 Aan: Robbie Joostenmailto:robbie_joos...@hotmail.com; CCP4BB@JISCMAIL.AC.UKmailto:CCP4BB@JISCMAIL.AC.UK Onderwerp: RE: [ccp4bb] paired refinement You need unmerged data to calculate cc1/2. That's not the sort of data you get from the PDB. Yes, good point; I forgot about that. But anyway, we have a fairly simple automated test that we can use on a case-by-case basis. I would argue that that is nicer than a empirical cut-off that may or may not be correct for you case. An automated test for what? And is it implemented somewhere convenient? JPK Cheers, Robbie Sent with my Windows Phone Van: Keller, Jacobmailto:kell...@janelia.hhmi.org Verzonden: 2-7-2015 20:12 Aan: CCP4BB@JISCMAIL.AC.UKmailto:CCP4BB@JISCMAIL.AC.UK Onderwerp: Re: [ccp4bb] paired refinement Well, in that case, one could simply look at the plot of CC1/2 versus resolution and see the step up to one, conclude something was off. I wonder whether PDB REDO was able to get some empirically-determined values for CC1/2 cutoffs by comparing paired refinement versus CC1/2 versus other parameters? JPK -Original Message- From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Dale Tronrud Sent: Thursday, July 02, 2015 1:46 PM To: CCP4BB@JISCMAIL.AC.UKmailto:CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] paired refinement While I was puzzling over an entry in the PDB some years ago (since obsoleted) I noticed that all the high resolution amplitudes were equal to 11.0! This was before CC1/2 but for this structure it would have been equal to one, and yet the outer data were useless. A practical test like paired refinement can't be fooled in this way. Dale Tronrud On 7/2/2015 10:25 AM, Edward A. Berry wrote: My take on this- No one has been willing to specify a cutoff (and probably there is no rigorous way to mathematically define the cutoff) and say If CC* (or CCfree or whatever) is below X then it will not improve your structure, if above X then it will. Probably depends among other things on how strong the lower resolution data is, how good the structure is without the added data. On the other hand in paired refinement, if adding the data improves the structure as measured by Rfree in a zone excluding the added data, then it is hard to deny that that data are worth including. eab On 07/02/2015 12:52 PM, Keller, Jacob wrote: Wasn’t all of this put to bed through the implementation of CC measures? JPK *From:*CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] *On Behalf Of *Robbie Joosten *Sent:* Thursday, July 02, 2015 12:46 PM *To:* CCP4BB@JISCMAIL.AC.UKmailto:CCP4BB@JISCMAIL.AC.UK *Subject:* Re: [ccp4bb] paired refinement But it is not the R-free of the shell here. In paired refinement you take the R-free of the reflections outside the shell. Cheers, Robbie Sent with my Windows Phone - - - - - - - - - - - - - - - --- *Van: *Edward A. Berry mailto:ber...@upstate.edu *Verzonden: *2-7-2015 18:43 *Aan: *CCP4BB@JISCMAIL.AC.UKmailto:*CCP4BB@JISCMAIL.AC.UK mailto:CCP4BB@JISCMAIL.AC.UK *Onderwerp: *Re: [ccp4bb] paired refinement Another criterion for cutoff, also requiring the structure to be solved, is the agreement between data and structure, e.g. Rfree or CCfree. I think it is very unlikely that you could get Rfree =.2493 in a shell which contains only noise. So I would suggest doing paired refinement to 2.2 and 2.1 A (if the data is available). On 07/01/2015 07:15 PM, Eric Karg wrote: Hi all, I have a dataset processed in XDS to 2.3 A (based on CC1/2). I'm trying to do paired refinement to determine the optimal
Re: [ccp4bb] Heavy-atom derivatives
Dear Giulliana, I imagine this may well be of assistance:- *J. Appl. Cryst.* (2009). *42*, 540-544[ doi:10.1107/S0021889809012370 http://dx.doi.org/10.1107/S0021889809012370 ] HATODAS II - heavy-atom database system with potentiality scoringM. Sugahara http://scripts.iucr.org/cgi-bin/citedin?search_on=nameauthor_name=Sugahara,%20M., Y. Asada http://scripts.iucr.org/cgi-bin/citedin?search_on=nameauthor_name=Asada,%20Y., H. Shimada http://scripts.iucr.org/cgi-bin/citedin?search_on=nameauthor_name=Shimada,%20H., H. Taka http://scripts.iucr.org/cgi-bin/citedin?search_on=nameauthor_name=Taka,%20H. and N. Kunishima http://scripts.iucr.org/cgi-bin/citedin?search_on=nameauthor_name=Kunishima,%20N. Best wishes, John On Mon, Jun 29, 2015 at 1:47 PM, Giulliana Rangel giulliana.ran...@gmail.com wrote: Dear all, I'm looking for a method to solve the phase problem. Thus, I would like some help about heavy-atom derivatives. What is the best heavy-atom to make soaking? I tried Iodine (because my protein have a lot of tyrosines), but no results... I read some papers about quick-soak, its a good method? Using platine, Hg, Br... Any suggestions? Thank you so much. Best wishes, -- Giulliana Rangel M.S. Biotechnology Program - UNIFESP Structural Biology Laboratory Phone: +55 (12) 3309-9698 330 Talim St ZIP CODE 12231-280 São José dos Campos - SP/ BR -- Professor John R Helliwell DSc
Re: [ccp4bb] Rfree below Rwork
Dear Smith, when you expand to P1, pointless should suggest the space group you expanded from, unless you fiddled with the data after expansion. Regards, Tim On 07/01/2015 04:43 AM, Smith Liu wrote: If both the PDB and mtz for the pdb have been assigned to P1 space group for some reason, can this lead to Rwork higher than Rfree during refinement? If after converting my PDB and mtz to P1 space group, and I have forgotten what is the original space group for my PDB and mtz before conversion to P1 space group, is any method which can recover the original space group for my PDBand mtz, so that in the following refine Rwork would be lower than Rfree? Smith At 2015-07-01 01:55:22, Eleanor Dodson eleanor.dod...@york.ac.uk wrote: I suppose if I was the referee for this structure and your FreeR is so close to the Rfactor I would ask you to ensure you had the right space group - is the 6 fold NCS actually 2 fold NCS with a crystallographic 3 fold.. Cases occur where R32 is indexed as C2.. Certainly if the Rfree set is assigned randomly to reflections which are symmetry equivalents then you see this phenomena of Rfree = Rfactor Eleanor On 30 June 2015 at 18:26, Gerard Bricogne g...@globalphasing.com wrote: Dear Wolfram, I have a perhaps optimistic view of the effect of high-order NCS on Rfree, in the sense that I don't view it as a problem. People have agonised to extreme degrees over the difficulty of choosing a free set of reflections that would produce the expected gap between Rwork and Rfree, and some of the conclusions were that you would need to hide almost half of your data in some cases! I think it is best to remember that the idea of cross-validation by Rfree is to prevent overfitting, i.e. ending up with a model that fits the amplitudes too well compared to how well it determines the phases. In the case of high-order NCS (in your case, the U/V ratio that the old papers on NCS identified as the key quantity to measure the phasing power of NCS would be less than 0.1!) the phases and the amplitudes are so tightly coupled that it is simply impossible to fit the amplitudes without delivering phases of an equally good quality. In other words there is no overfitting problem (provided you do have good and complete data) and the difference between Rfree and Rwork is simply within the bounds of the statistical spread of Rfree depending on the free set chosen. You are lucky to have 6-fold NCS, so don't let any reviewer convince you that it is a curse, and make you suffer for it :-) . With best wishes, Gerard. -- On Tue, Jun 30, 2015 at 12:58:44PM -0400, wtempel wrote: Hello, my question concerns refinement of a structure with 6-fold NCS (local automatic restraints in REFMAC) against 2.8 A data. The size of my free set is 1172 selected in thin resolution shells (SFTOOLS) and corresponding to 4.3 % of reflections. A refmac run of 10 cycles of TLS and 10 cycles of CGMAT starts out at Rfree/Rcryst 0.271/0.272. After the 10th TLS cycle I have 0.227/0.224. Yes, Rfree Rcryst. At the end of CGMAT I have 0.2072/0.2071. I understand that NCS stresses the independence assumption of the free set. Am I correct in believing that Rfree *may* be smaller than Rcryst even in the absence of a major mistake? My hope is that the combined wisdom of ccp4bb followers can point out my possible mistake, suggest tests that I may perform to avoid them and, possibly, arguments in defense of a crystallographic model with Rfree Rcryst. Many thanks, Wolfram Tempel -- === * * * Gerard Bricogne g...@globalphasing.com * * * * Global Phasing Ltd. * * Sheraton House, Castle Park Tel: +44-(0)1223-353033 * * Cambridge CB3 0AX, UK Fax: +44-(0)1223-366889 * * * === -- -- Dr Tim Gruene Institut fuer anorganische Chemie Tammannstr. 4 D-37077 Goettingen phone: +49 (0)551 39 22149 GPG Key ID = A46BEE1A signature.asc Description: OpenPGP digital signature
Re: [ccp4bb] paired refinement
Hi Robbie, I have been wondering how much information would be present in a weighted CC1/2 with weights from the ML refinement program. As I understand the concept behind paired refinement, one can use much higher resolution data in refinement than you would expect from the (classical) scaling statistics, because refinement programs down-weight the outliers. Maybe a weighted CC1/2 would be a convenient short cut the more time consuming paired refinemement? How easily could the calculation of a weighted CC1/2 be implemented in refmac? Refmac would have to merge the data internally, but that's just a simple formula to implement. Best, Tim On 07/02/2015 08:28 PM, Robbie Joosten wrote: Hi Jacob, You need unmerged data to calculate cc1/2. That's not the sort of data you get from the PDB. But anyway, we have a fairly simple automated test that we can use on a case-by-case basis. I would argue that that is nicer than a empirical cut-off that may or may not be correct for you case. Cheers, Robbie Sent with my Windows Phone Van: Keller, Jacobmailto:kell...@janelia.hhmi.org Verzonden: 2-7-2015 20:12 Aan: CCP4BB@JISCMAIL.AC.UKmailto:CCP4BB@JISCMAIL.AC.UK Onderwerp: Re: [ccp4bb] paired refinement Well, in that case, one could simply look at the plot of CC1/2 versus resolution and see the step up to one, conclude something was off. I wonder whether PDB REDO was able to get some empirically-determined values for CC1/2 cutoffs by comparing paired refinement versus CC1/2 versus other parameters? JPK -Original Message- From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Dale Tronrud Sent: Thursday, July 02, 2015 1:46 PM To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] paired refinement While I was puzzling over an entry in the PDB some years ago (since obsoleted) I noticed that all the high resolution amplitudes were equal to 11.0! This was before CC1/2 but for this structure it would have been equal to one, and yet the outer data were useless. A practical test like paired refinement can't be fooled in this way. Dale Tronrud On 7/2/2015 10:25 AM, Edward A. Berry wrote: My take on this- No one has been willing to specify a cutoff (and probably there is no rigorous way to mathematically define the cutoff) and say If CC* (or CCfree or whatever) is below X then it will not improve your structure, if above X then it will. Probably depends among other things on how strong the lower resolution data is, how good the structure is without the added data. On the other hand in paired refinement, if adding the data improves the structure as measured by Rfree in a zone excluding the added data, then it is hard to deny that that data are worth including. eab On 07/02/2015 12:52 PM, Keller, Jacob wrote: Wasn’t all of this put to bed through the implementation of CC measures? JPK *From:*CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] *On Behalf Of *Robbie Joosten *Sent:* Thursday, July 02, 2015 12:46 PM *To:* CCP4BB@JISCMAIL.AC.UK *Subject:* Re: [ccp4bb] paired refinement But it is not the R-free of the shell here. In paired refinement you take the R-free of the reflections outside the shell. Cheers, Robbie Sent with my Windows Phone - - - - - - - - - - - - - - - --- *Van: *Edward A. Berry mailto:ber...@upstate.edu *Verzonden: *2-7-2015 18:43 *Aan: *CCP4BB@JISCMAIL.AC.UK mailto:CCP4BB@JISCMAIL.AC.UK *Onderwerp: *Re: [ccp4bb] paired refinement Another criterion for cutoff, also requiring the structure to be solved, is the agreement between data and structure, e.g. Rfree or CCfree. I think it is very unlikely that you could get Rfree =.2493 in a shell which contains only noise. So I would suggest doing paired refinement to 2.2 and 2.1 A (if the data is available). On 07/01/2015 07:15 PM, Eric Karg wrote: Hi all, I have a dataset processed in XDS to 2.3 A (based on CC1/2). I'm
[ccp4bb] Nickel coordination
Hello again. How does one prepare restraints for a Ni ion with four ligands: - NE2 atoms from two His residues - N and O from one Gly residue REFMAC nailed down the distance. How do I specify a near square-planar arrangement of the ligands? Thank you, Wolfram Tempel
