[ccp4bb] Instruct-NL Workshop on Wednesday April 25, 2012, in Leiden, the Netherlands
Dear Colleague, INSTRUCT-NL organize a one-day Instruct Workshop which will be on Wednesday April 25, 2012, in Leiden, the Netherlands, with as theme Developments in Structural Biology: Instruct meets industry The program can be found at http://www.structuralbiology.eu/support/whats-on/calendar-events/developments-structural-biology-instruct-meets-industry. Confirmed speakers include Dave Stuart (Oxford), Bram Koster (LUMC), Elspeth Gordon (ESRF), Darren Hart (EMBL), Raimond Ravelli (LUMC), Imre Berger (ESRF), Jack Johnson (The Scripps Research Institute), Albert Heck (Utrecht University), Marc Baldus (Utrecht University) and Clemens Löwik (LUMC). During these talks, techniques including protein production, proteomics, mass spectrometry, solid state NMR, X-ray crystallography, electron microscopy, and more, will be discussed. We welcome scientists interested in the field of Structural Biology, especially those interested in considering to use, or offer access to, technological infrastructures on a collaborative scientific base or from a commercial industrial point of view. For more information on how Instruct can be of help to you, please check the Instruct website at http://www.structuralbiology.eu/ . The workshop will be held at Naturalis, the Dutch national museum for natural history (http://www.naturalis.nl/en/visit-us/getting-there/). The venue is conveniently located a few minutes walking distance from Leiden Central Railway Station, which is only a 20 minute train ride from Amsterdam Airport Schiphol. The venue is also a mere 15 minute walk from the novel Netherlands Center of Electron Nanoscopy (NeCEN, http://www.necen.nl ). Please register at http://www.structuralbiology.eu/content/registration-developments-structural-biology-instruct-meets-industry The deadline for registration is the 18th of April 2012. Please distribute this mail to people you think might be interested. Best regards, Bram Koster: a.j.kos...@lumc.nl Albert Heck: a.j.r.h...@uu.nl Esther van Duijn: e.vandu...@uu.nl Raimond Ravelli: rave...@lumc.nl Reinout Raijmakers: r.raijmak...@uu.nl
Re: [ccp4bb] Refmac executables - win vs linux in RHEL VM
On Sat, Apr 07, 2012 at 08:42:47AM -0700, Bernhard Rupp (Hofkristallrat a.D.) wrote: Something the developers might be interested in: The Refmac_5.6.0117 32-bit windows binaries run native on a win64 3-4x slower than those from the linux distribution run Thanks for benchmarking. If you'd like to test refmac 5.7 that will be included in CCP4 6.3 I can make pre-release binaries later this week and put it on ftp. (I suppose it won't be slower than Linux version, but we'll see.) One thing that slows down *both* Linux and Windows version is the GFORTRAN_UNBUFFERED_ALL env. variable. It was set as a temporary workaround in ver. 6.2, but has been already removed from the coming release. If you unset it for refmac from 6.2 you can get a few lines in the output (in terminal) in wrong order, but the program should run notably faster. Most peculiaralthough I think but I do not know whether the linux binaries are 64 bit If it's from CCP4 it's 32-bit. You can check it by typing file /path/to/binary. You'll get in the output either ELF 32-bit LSB executable or ELF 64-bit LSB executable Marcin
[ccp4bb] Disorder or poor phases?
Hi all, Assume that the diffraction resolution is low (say 3.0A or worse) and the model (a high resolution homologue, from 2A xray data is available) was docked into experimental phases (say 4A or worse) and extended to the 3.0A data using refinement (the high resolution model as a source of restraints). There are some conformational differences between the high resolution model and the target crystal. The author observes that in the 2fofc map at 3A, most of the model shows reasonable density, but for a stretch of backbone the density is weak. Is the weakness of the density in this region because of disorder or bad model phases? Would love people's thoughts on this one, F - Francis E. Reyes M.Sc. 215 UCB University of Colorado at Boulder
Re: [ccp4bb] Disorder or poor phases?
-BEGIN PGP SIGNED MESSAGE- Hash: SHA1 Dear Francis, the phases calculated from the model affect the whole unit cell hence it is more likely this is real(-space, local) disorder rather than poor phases. Regards, Tim P.S.: The author should not look at an 2fofc-map but a sigma-A-weighted map to reduce model bias. On 04/10/12 17:22, Francis E Reyes wrote: Hi all, Assume that the diffraction resolution is low (say 3.0A or worse) and the model (a high resolution homologue, from 2A xray data is available) was docked into experimental phases (say 4A or worse) and extended to the 3.0A data using refinement (the high resolution model as a source of restraints). There are some conformational differences between the high resolution model and the target crystal. The author observes that in the 2fofc map at 3A, most of the model shows reasonable density, but for a stretch of backbone the density is weak. Is the weakness of the density in this region because of disorder or bad model phases? Would love people's thoughts on this one, F - Francis E. Reyes M.Sc. 215 UCB University of Colorado at Boulder - -- - -- Dr Tim Gruene Institut fuer anorganische Chemie Tammannstr. 4 D-37077 Goettingen GPG Key ID = A46BEE1A -BEGIN PGP SIGNATURE- Version: GnuPG v1.4.12 (GNU/Linux) Comment: Using GnuPG with Mozilla - http://enigmail.mozdev.org/ iD8DBQFPhFP3UxlJ7aRr7hoRAj4HAKDpHCsN+tBKhDAcOYmIe5c58ThG+gCeMujG pAJxRNuJHE4+oFRPSYx4bnc= =s3uw -END PGP SIGNATURE-
Re: [ccp4bb] Disorder or poor phases?
Dale Thank you for the case study. I will certainly remember it when I next see: I don't see density for these atoms therefore they must be disordered. You do mention though, that when you were able to assign the sequence to the beta sheets, that the loop regions became clear. I consider the case (which a majority of cases seem to be), where the author has built and sequence assigned 95% of the ASU, but is unable to model a loop region. One possibility is that the loop is truly disordered (95% of the ASU is built and is presumably right), the other possibility is that there's an inherent error in the existing structure that is affecting the interpretation of the loop region. The errors are probably extremely subtle and distributed throughout the model (think of the improvements DEN refinement gave for the rerefinement of p97). I guess in either case, because of the dependency of the map on the existing set of phases it's difficult to determine whether it's truly disordered or not. P.S.: The author should not look at an 2fofc-map but a sigma-A-weighted map to reduce model bias. Tim, I assume a sigmaA weighted 2Fo-Fc map (which I believe is the default for most crystallographic refinement packages). F - Francis E. Reyes M.Sc. 215 UCB University of Colorado at Boulder
Re: [ccp4bb] Disorder or poor phases?
Dear Dale, There is perhaps a third factor in the progress you were able to make, namely the improvement in the refinement programs. Your first results were probably obtained with a least-squares-based program, while the more recent would have come from maximum-likelihood-based ones. The difference lies in the quality of the phase information produced from the model through comparison of Fo and Fc, with much greater bias-correction capabilities in the ML approach. Here, it removed the bias towards some regions being absent in the model, and made them no longer be absent in the maps. So it is a question of the quality of the phase information. With best wishes, Gerard. -- On Tue, Apr 10, 2012 at 12:00:28PM -0700, Dale Tronrud wrote: The phases do have effects all over the unit cell but that does not prevent them from constructively and destructively interfering with one another in particular locations. Some years ago I refined a model of the bacteriochlorophyll containing protein to a 1.9 A data set when the sequence of that protein was unknown. This is primarily a beta sheet protein and a number of the loops between the strands were disordered. Later the amino acid sequence was determined and I finished the refinement after building in these corrections. The same data set was used, but a number of the loops had become ordered. While the earlier model (3BCL) had 357 amino acids the final model (4BCL) had 366. These nine amino acids didn't become ordered over the intervening years. They were just as ordered when I was building w/o a sequence, it is just that I couldn't see how to build them based on the map's appearance. One possibility is that the density for these residues was weak and the noise (that was uniform over the entire map) obliterated their signal where it only obscured the stronger density. Another possibility is that the better model had a better match of the low resolution F's and less intense ripples radiating from the surface of the molecule, resulting in things sticking out being less affected. Whatever the details, the density for these amino acids were too weak to model with the poorer model phases and became buildable with better phases. The fact that they could not be seen in the early map was not an indication that they were disordered. The first six amino acids of this protein have never been seen in any map, including the 1.3 A resolution model 3EOJ (which by all rights should have been called 5BCL ;-) ). These residues appear to be truly disordered. Going back to 3BCL - The map for this model is missing density for a number of residues of which we know some are disordered and some simply unmodelable because of the low quality of the phases. I don't know of a way, looking at that map alone, of deciding which is which. Because of this observation I don't believe it is supportable to say I don't see density for these atoms therefore they must be disordered. Additional evidence is required. Dale Tronrud On 04/10/12 08:38, Tim Gruene wrote: Dear Francis, the phases calculated from the model affect the whole unit cell hence it is more likely this is real(-space, local) disorder rather than poor phases. Regards, Tim P.S.: The author should not look at an 2fofc-map but a sigma-A-weighted map to reduce model bias. On 04/10/12 17:22, Francis E Reyes wrote: Hi all, Assume that the diffraction resolution is low (say 3.0A or worse) and the model (a high resolution homologue, from 2A xray data is available) was docked into experimental phases (say 4A or worse) and extended to the 3.0A data using refinement (the high resolution model as a source of restraints). There are some conformational differences between the high resolution model and the target crystal. The author observes that in the 2fofc map at 3A, most of the model shows reasonable density, but for a stretch of backbone the density is weak. Is the weakness of the density in this region because of disorder or bad model phases? Would love people's thoughts on this one, F - Francis E. Reyes M.Sc. 215 UCB University of Colorado at Boulder
Re: [ccp4bb] Disorder or poor phases?
Dear Gerard, No, the updated model (4BCL) was published in 1993 (although apparently not deposited until 1998 - What was wrong with me?) Both were refined with that classic least-squares program TNT. I hope there was some improvement in the software between 1986 and 1993, and I always tried to work with the most recent version, but there wasn't a switch in target function. I agree that the distortions in these maps would have been less if an ML approach had been used and perhaps the location of the disordered residues would have been apparent earlier in the process. Maybe this sort of problem will not be seen again at 1.9 A resolution. My goal was simply to provide an example where errors due to model phases didn't distribute evenly throughout the map but had greater consequence in some locations. Dale On 04/10/12 13:45, Gerard Bricogne wrote: Dear Dale, There is perhaps a third factor in the progress you were able to make, namely the improvement in the refinement programs. Your first results were probably obtained with a least-squares-based program, while the more recent would have come from maximum-likelihood-based ones. The difference lies in the quality of the phase information produced from the model through comparison of Fo and Fc, with much greater bias-correction capabilities in the ML approach. Here, it removed the bias towards some regions being absent in the model, and made them no longer be absent in the maps. So it is a question of the quality of the phase information. With best wishes, Gerard. -- On Tue, Apr 10, 2012 at 12:00:28PM -0700, Dale Tronrud wrote: The phases do have effects all over the unit cell but that does not prevent them from constructively and destructively interfering with one another in particular locations. Some years ago I refined a model of the bacteriochlorophyll containing protein to a 1.9 A data set when the sequence of that protein was unknown. This is primarily a beta sheet protein and a number of the loops between the strands were disordered. Later the amino acid sequence was determined and I finished the refinement after building in these corrections. The same data set was used, but a number of the loops had become ordered. While the earlier model (3BCL) had 357 amino acids the final model (4BCL) had 366. These nine amino acids didn't become ordered over the intervening years. They were just as ordered when I was building w/o a sequence, it is just that I couldn't see how to build them based on the map's appearance. One possibility is that the density for these residues was weak and the noise (that was uniform over the entire map) obliterated their signal where it only obscured the stronger density. Another possibility is that the better model had a better match of the low resolution F's and less intense ripples radiating from the surface of the molecule, resulting in things sticking out being less affected. Whatever the details, the density for these amino acids were too weak to model with the poorer model phases and became buildable with better phases. The fact that they could not be seen in the early map was not an indication that they were disordered. The first six amino acids of this protein have never been seen in any map, including the 1.3 A resolution model 3EOJ (which by all rights should have been called 5BCL ;-) ). These residues appear to be truly disordered. Going back to 3BCL - The map for this model is missing density for a number of residues of which we know some are disordered and some simply unmodelable because of the low quality of the phases. I don't know of a way, looking at that map alone, of deciding which is which. Because of this observation I don't believe it is supportable to say I don't see density for these atoms therefore they must be disordered. Additional evidence is required. Dale Tronrud On 04/10/12 08:38, Tim Gruene wrote: Dear Francis, the phases calculated from the model affect the whole unit cell hence it is more likely this is real(-space, local) disorder rather than poor phases. Regards, Tim P.S.: The author should not look at an 2fofc-map but a sigma-A-weighted map to reduce model bias. On 04/10/12 17:22, Francis E Reyes wrote: Hi all, Assume that the diffraction resolution is low (say 3.0A or worse) and the model (a high resolution homologue, from 2A xray data is available) was docked into experimental phases (say 4A or worse) and extended to the 3.0A data using refinement (the high resolution model as a source of restraints). There are some conformational differences between the high resolution model and the target crystal. The author observes that in the 2fofc map at 3A, most of the model shows reasonable density, but for a stretch of backbone the density is weak. Is the weakness of the density in this region because of disorder or bad model phases? Would
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Re: [ccp4bb] Disorder or poor phases?
Hi Dale, my experience is that high-B regions may become visible in maps only late in refinement. So my answer to the original poster would be - both global reciprocal-space (phase quality) and local real-space (high mobility) features contribute to a region not appearing ordered in the map. This would be supported by your experience if those residues that you could not model in 3BCL had high (or at least higher) B-factors compared to the rest of the model. Is that so? best, Kay
