Re: [ccp4bb] a challenge
I am absolutely delighted at the response I have gotten to my little John Henry Challenge! Three people already have managed to do the impossible. Congratulations to George Sheldrick, Pavol Skubak and Raj Pannu for finding ways to improve the phases over the ones I originally obtained (using the default settings of mlphare and dm) and build their way out of it. This is quite useful information! At least it is to me. Nevertheless, I do think Frances Reyes has a point. This was meant to be a map interpretation challenge, and not a SAD-phasing challenge. I appreciate that the two are linked, but the reason I did not initially provide the anomalous data is because I thought it would be too much to ask people to re-do all the phasing, etc. Yes, there do appear to be ways to improve the maps beyond the particular way I phased them, but no matter how good your phasing program is, there will always be a level of anomalous signal that will lead to phases that are off enough to make building the model impossible. Basically, once the map gets bad enough that just as many wrong atoms get built in as right atoms, then there is no escape. However, I think human beings should still have an advantage when it comes to pattern recognition, and I remain curious to see if an insightful crystallographer can tip that balance in the right direction. I am also still curious to see if tweaking some setting on some automated building program will do that too. So, my original question remains: are automated building programs better than humans? Any human? I therefore declare the John Henry Challenge still open. But yes, improving the phases can tip the balance too, and the accuracy of the anomalous differences will ultimately affect the accuracy of the phases, and so on. This is a much broader challenge. And I think the best way to frame it is with the question: How low can the anomalous signal be before any conceivable approach fails? and perhaps: What is the best procedure to use for weak anomalous signal? For those who are interested in joining George, Pavol, Raj and others in this new challenge, the full spectrum of difficulty from trivial (100% Se incorporation) to a complete waste of time (0% Se, 100% S) is here: http://bl831.als.lbl.gov/~jamesh/challenge/occ_scan/ The impossible.mtz for the John Henry Map Interpretation Challenge was derived from frac0.79.mtz and possible.mtz from frac0.78.mtz. These simulated 31% and 32% Se incorporation into Met side chains (respectively). It has now been shown that both of these can be solved automatically if you do the phasing right. But what about frac0.80.mtz? Or frac0.90.mtz ? At least on this one coordinate of Se incorporation, the prowess of a particular approach can be given a score. For example, a score of 0.78 means that the indicated procedure could solve the frac0.78.mtz dataset, but not the frac0.79.mtz dataset. Based on the reports I have gotten back so far, the difficulty score lineup is: score method 0.86 xds, xscale, right sites, crank2 (Pavol Skubak) 0.78 xds, xscale, right sites, mlphare, dm, phenix.autobuild using 20 models (James Holton) 0.75 xds, xscale, right sites, mlphare, dm, buccaneer/refmac/dm (James Holton) 0.71 xds, xscale, right sites, mlphare, dm, ARP/wARP 7.3 (James Holton) 0.51 xds, xscale, right sites, mlphare, dm, ARP/wARP 6.1.1 (James Holton) Note that all of these attempts cheated on the sites. Finding the sites seems to be harder than solving the structure once you've got them. That lineup is: score method 0.82 cheating: xds, xscale, right phases, anomalous difference Fourier (James Holton) 0.79 xds, xscale, shelxc/d/e 3.5A NTRY=1 (George Sheldrick) 0.74 xds, autorickshaw (Santosh Panjikar) 0.65xds, xscale, phenix.hyss --search=full (James Holton) 0.60 xds, xscale, shelxc/d with NTRY=100 (James Holton) Where again the score is the dataset where the heavy atom site constellation found is close enough to the right one to move forward. This transition, like the model-building one, is remarkably sharp, particularly if you let each step run for a lot of cycles. The graph for model-building is here: http://bl831.als.lbl.gov/~jamesh/challenge/build_CC_vs_frac.png Note how the final map quality is pretty much independent of the initial map quality, up to the point where it all goes wrong. I think this again is an example of the solution needing to be at least half right before it can be improved. But perhaps someone can prove me wrong on that one? For those who want the unmerged data, I have all the XDS_ASCII.HKL files here: http://bl831.als.lbl.gov/~jamesh/challenge/occ_scan/XDS_ASCII.tgz If you'd like to go all the way back to the images, you can get them from here: http://bl831.als.lbl.gov/~jamesh/workshop2/ the badsignal dataset is what produced frac1.00.mtz, and goodsignal produced frac0.00.mtz. You can generate anything in
[ccp4bb] ccp4 update
Dear CCP4 maintainers, I've come to appreciate the CCP4 update functionality, which, in our multiuser network (RHEL 6.2), I used to invoke by calling $CCP4/bin/update. Update 012 removed that script with no immediately obvious replacement. Was that on purpose? Is there a way of updating CCP4 from the command line without calling CCP4i? Thanks Andreas (from update.log: [Thu Jan 3 2013 11:00:21] Ready to make changes --- applying update 6.3.0-012 --- update header read --- creating restore package, please wait ... --- done ... file '/csb/soft/Linux64/share/ccp4-6.3.0/bin/update' removed ... file '/csb/soft/Linux64/share/ccp4-6.3.0/lib_exec/update' removed ... file '/csb/soft/Linux64/share/ccp4-6.3.0/lib_exec/_update' removed ... directory '/csb/soft/Linux64/share/ccp4-6.3.0/lib_exec' removed some more blah blah) -- Andreas Förster, Research Associate Paul Freemont Xiaodong Zhang Labs Department of Biochemistry, Imperial College London http://www.msf.bio.ic.ac.uk
[ccp4bb] Post-doc positions @Amsterdam
Post-doctoral positions are available in the Netherlands Cancer Institute in the groups of Anastassis (Tassos) Perrakis and Titia Sixma (http://xtal.nki.nl). The Netherlands Cancer Institute (http://www.nki.nl) is a center of excellence with a high standard of biological research and an interactive international atmosphere. It is located in Amsterdam, with all its cultural amenities, close to the Schiphol airport. The groups are closely associated and have an interest in structural studies coupled to functional analysis, combined with activities in method development for structural biology. Our department also hosts the Protein Facility with equipment that includes high throughput crystallization and crystal visualization robotics, an X-ray facility, Biacore surface plasmon resonance, isothermal titration calorimetry (ITC), static light scattering (MALLS), versatile systems for fluorescence based equilibrium and transient state kinetics analysis (a versatile plate reader and stopped-flow), an Orbitrap mass spec, and operational facilities for E.coli, insect cells and mammalian cells based protein expression. The project in the laboratory of Titia Sixma is focused at the interface of ubiquitin (de)conjugation and DNA repair. The projects in Tassos Perrakis group are about Autotaxin and/or proteins that regulate mitotic progression. We are looking for enthusiastic researchers with experience in molecular biology and/or biochemistry and/or protein crystallography. Applicants should write an e-mail with CV and names of three references to t.si...@nki.nl or a.perra...@nki.nl Tassos Titia
Re: [ccp4bb] a challenge
What is the best procedure to use for weak anomalous signal That opens up the can of worms which I'm happy to jump into. We've had very good success in the years 2003-2009 with shelx for finding sites (sometimes more than 1 trials) then force feeding them to sharp for phase improvement. We should also say most of the times in particular in the more difficult cases xds made the difference in detectable anomalous signal. And no we still have not published this. With we I mean Marc Robien and myself during our SGPP times. Jürgen .. Jürgen Bosch Johns Hopkins Bloomberg School of Public Health Department of Biochemistry Molecular Biology Johns Hopkins Malaria Research Institute 615 North Wolfe Street, W8708 Baltimore, MD 21205 Phone: +1-410-614-4742 Lab: +1-410-614-4894 Fax: +1-410-955-3655 http://lupo.jhsph.edu On Jan 14, 2013, at 3:13, James Holton jmhol...@lbl.gov wrote: What is the best procedure to use for weak anomalous signal
[ccp4bb] Two Postdoc positions at the University of Oxford, SGC
Dear all, We are seeking two post-doc scientists in the Metabolic Rare Diseases group at the Structural Genomics Consortium (SGC), University of Oxford: 1. *Protein crystallographer*, driving multiple gene-to-structure projects to understand inborn errors of metabolism: https://www.recruit.ox.ac.uk/pls/hrisliverecruit/erq_jobspec_version_4.jobspec?p_id=106057 2. Industry-funded *Protein biochemist/structural biologist*, working on mechanism and small molecule development for protein misfolding disorders: https://www.recruit.ox.ac.uk/pls/hrisliverecruit/erq_jobspec_version_4.jobspec?p_id=106055 The successful candidates will - benefit from the SGC high-throughput platform for structural/biochemical characterization - work closely with an established network of geneticists, clinicians and drug developers - have regular access to the nearby Diamond synchrotron for data collection Visit the SGC Metabolic Rare Diseases group: http://www.thesgc.org/wyatt Informal enquiries to: wyatt@sgc.ox.ac.uk Application deadline:* Noon 7th Feb 2013 (GMT)* * * * * Dr Wyatt W. Yue Principal Investigator, Metabolic Rare Diseases Structural Genomics Consortium University of Oxford UK OX3 7DQ +44 (0)1865 617757 http://www.thesgc.org/wyatt
[ccp4bb] engh huber
To what extent modern geometric restraints have been upgraded over original EnghHuber? And where I can find a consensus set of values (with variances)? For example, Fisher et al., Acta D68:800 discusses how histidine angles change with protonation, and refers to EnghHuber when it says that ND1-CE1-NE2 goes from 111.2 to 107.5 when histidine acquires positive charge (Fig.6). But angle table (Table 3) in original EnghHuber from 1991 does not have any 107.5 value and seems to suggest that the numbers should rather be 111.7+-1.3 and 108.4+-1.0, respectively. I understand that these values are derived from structural databases and thus can be frequently updated. Is there some resource where most current values would be listed? Cheers, Ed. -- After much deep and profound brain things inside my head, I have decided to thank you for bringing peace to our home. Julian, King of Lemurs
Re: [ccp4bb] engh huber
Hi Ed, Chapter 18.3 of international tables vol F includes values designated EH99 which are from a more recent CSD release than the original 1991 Engh Huber paper. R. A. Engh and R. Huber. Structure quality and target parameters. International Tables for Crystallography (2012). Vol. F, ch. 18.3, pp. 474-484 doi: 10.1107/9780955360206857 http://it.iucr.org/Fb/ch18o3v0001/ Also, the Buster groups' Grade server provides dynamic use of the CSD database to derive restraints. http://grade.globalphasing.org And the PURY restraint database has restraints derived from recent CSD releases. I belive it requires a current CSD license is required for use. http://pury.ijs.si/ Regards, Mitch -Original Message- From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Ed Pozharski Sent: Monday, January 14, 2013 9:55 AM To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] engh huber To what extent modern geometric restraints have been upgraded over original EnghHuber? And where I can find a consensus set of values (with variances)? For example, Fisher et al., Acta D68:800 discusses how histidine angles change with protonation, and refers to EnghHuber when it says that ND1-CE1-NE2 goes from 111.2 to 107.5 when histidine acquires positive charge (Fig.6). But angle table (Table 3) in original EnghHuber from 1991 does not have any 107.5 value and seems to suggest that the numbers should rather be 111.7+-1.3 and 108.4+-1.0, respectively. I understand that these values are derived from structural databases and thus can be frequently updated. Is there some resource where most current values would be listed? Cheers, Ed. -- After much deep and profound brain things inside my head, I have decided to thank you for bringing peace to our home. Julian, King of Lemurs
Re: [ccp4bb] engh huber
There was an update by EH in 2001 in the International Tables Vol F. There are a small number of modifications to the 1991 values in the update as well as the addition of several conformational variabilities. If I understand correctly, Refmac and Phenix use the 2001 values, with the only conformational variability being some changes with cis-peptide bonds. Shelxl still uses EH 1991. Dale Tronrud On 01/14/13 09:54, Ed Pozharski wrote: To what extent modern geometric restraints have been upgraded over original EnghHuber? And where I can find a consensus set of values (with variances)? For example, Fisher et al., Acta D68:800 discusses how histidine angles change with protonation, and refers to EnghHuber when it says that ND1-CE1-NE2 goes from 111.2 to 107.5 when histidine acquires positive charge (Fig.6). But angle table (Table 3) in original EnghHuber from 1991 does not have any 107.5 value and seems to suggest that the numbers should rather be 111.7+-1.3 and 108.4+-1.0, respectively. I understand that these values are derived from structural databases and thus can be frequently updated. Is there some resource where most current values would be listed? Cheers, Ed.
Re: [ccp4bb] engh huber
Article in the Tables is the answer to my question about the latest EnghHuber parameters. These still don't match Fig.6 from Fisher, but I am OK with using Tables for my internal purposes. Thanks to Mitchell and Dale for prompt response. Cheers, Ed. -- After much deep and profound brain things inside my head, I have decided to thank you for bringing peace to our home. Julian, King of Lemurs
Re: [ccp4bb] a challenge
-BEGIN PGP SIGNED MESSAGE- Hash: SHA1 Hello James and all other contributors, I admit not having read all contributions to this thread. I understand the John Henry Challenge as whether there is an 'automated way of producing a model from impossible.mtz'. From looking at it and without having gone all the way to a PDB-file my feeling is one could without too much effort from the baton mode in e.g. coot. I guess this is not what you (and this thread) mean by 'automated' which leaves the impression that crystallographers have become quite spoiled children for this notion undermines how much effort and ingenuity the authors of programs like coot, O, mifit, frodo, etc, etc, pp - compared how models were prepared before this algorithms had been implemented, there is a lot of automation even in looking at the skeleton of a map! Cheers, Tim On 01/14/2013 10:12 AM, James Holton wrote: I am absolutely delighted at the response I have gotten to my little John Henry Challenge! Three people already have managed to do the impossible. Congratulations to George Sheldrick, Pavol Skubak and Raj Pannu for finding ways to improve the phases over the ones I originally obtained (using the default settings of mlphare and dm) and build their way out of it. This is quite useful information! At least it is to me. Nevertheless, I do think Frances Reyes has a point. This was meant to be a map interpretation challenge, and not a SAD-phasing challenge. I appreciate that the two are linked, but the reason I did not initially provide the anomalous data is because I thought it would be too much to ask people to re-do all the phasing, etc. Yes, there do appear to be ways to improve the maps beyond the particular way I phased them, but no matter how good your phasing program is, there will always be a level of anomalous signal that will lead to phases that are off enough to make building the model impossible. Basically, once the map gets bad enough that just as many wrong atoms get built in as right atoms, then there is no escape. However, I think human beings should still have an advantage when it comes to pattern recognition, and I remain curious to see if an insightful crystallographer can tip that balance in the right direction. I am also still curious to see if tweaking some setting on some automated building program will do that too. So, my original question remains: are automated building programs better than humans? Any human? I therefore declare the John Henry Challenge still open. But yes, improving the phases can tip the balance too, and the accuracy of the anomalous differences will ultimately affect the accuracy of the phases, and so on. This is a much broader challenge. And I think the best way to frame it is with the question: How low can the anomalous signal be before any conceivable approach fails? and perhaps: What is the best procedure to use for weak anomalous signal? For those who are interested in joining George, Pavol, Raj and others in this new challenge, the full spectrum of difficulty from trivial (100% Se incorporation) to a complete waste of time (0% Se, 100% S) is here: http://bl831.als.lbl.gov/~jamesh/challenge/occ_scan/ The impossible.mtz for the John Henry Map Interpretation Challenge was derived from frac0.79.mtz and possible.mtz from frac0.78.mtz. These simulated 31% and 32% Se incorporation into Met side chains (respectively). It has now been shown that both of these can be solved automatically if you do the phasing right. But what about frac0.80.mtz? Or frac0.90.mtz ? At least on this one coordinate of Se incorporation, the prowess of a particular approach can be given a score. For example, a score of 0.78 means that the indicated procedure could solve the frac0.78.mtz dataset, but not the frac0.79.mtz dataset. Based on the reports I have gotten back so far, the difficulty score lineup is: score method 0.86 xds, xscale, right sites, crank2 (Pavol Skubak) 0.78 xds, xscale, right sites, mlphare, dm, phenix.autobuild using 20 models (James Holton) 0.75 xds, xscale, right sites, mlphare, dm, buccaneer/refmac/dm (James Holton) 0.71 xds, xscale, right sites, mlphare, dm, ARP/wARP 7.3 (James Holton) 0.51 xds, xscale, right sites, mlphare, dm, ARP/wARP 6.1.1 (James Holton) Note that all of these attempts cheated on the sites. Finding the sites seems to be harder than solving the structure once you've got them. That lineup is: score method 0.82 cheating: xds, xscale, right phases, anomalous difference Fourier (James Holton) 0.79 xds, xscale, shelxc/d/e 3.5A NTRY=1 (George Sheldrick) 0.74 xds, autorickshaw (Santosh Panjikar) 0.65xds, xscale, phenix.hyss --search=full (James Holton) 0.60 xds, xscale, shelxc/d with NTRY=100 (James Holton) Where again the score is the dataset where the heavy atom site constellation found is close enough to the right one to move
Re: [ccp4bb] offtopic: steady state kinetics
Over the last 12 hours I've received a large number of responses, and I'd like to thank you all for the detailed and helpful responses. Clearly, I've been doing this incorrectly, however, I do have a number of experiments already done that hopefully I can salvage something out of. Now, at high concentrations of substrate I do get a hyperbolic curve, suggesting some sort of saturation occurring, and a half max at ~ 0.5-1mM. Now, although I haven't done the kinetics experiment properly, is there anything I can interpret from this data? As a reminder, my incorrectly done assay has been fixed concentration enzyme (~10uM) + varying substrate, 90 minute reaction, and a quench and detect with UV post. Again, thank you all for helping out a hapless grad student.
Re: [ccp4bb] a challenge
On Mon, Jan 14, 2013 at 11:18 AM, Tim Gruene t...@shelx.uni-ac.gwdg.de wrote: I admit not having read all contributions to this thread. I understand the John Henry Challenge as whether there is an 'automated way of producing a model from impossible.mtz'. From looking at it and without having gone all the way to a PDB-file my feeling is one could without too much effort from the baton mode in e.g. coot. This should be even more possible if one also uses existing knowledge about the expected structure of the protein: a kinase domain is quite distinctive. So, James, how much external information from homologous structures are we allowed to use? Running Phaser would certainly be cheating, but if I take (for instance) a 25% identical kinase structure, manually align it to the map and/or a partial model, and use that as a guide to manually rebuild the target model, does that meet the terms of the challenge? -Nat
[ccp4bb] Fwd: [ccp4bb] Annual CCP4 summer school in USA, at APS, June 19-26
Dear Colleagues, We are pleased to announce the sixth annual CCP4 summer school at Advanced Photon Source (APS), Argonne National Laboratory (ANL). All details can be found at http://www.ccp4.ac.uk/schools/APS-2013/index.php Title: CCP4 school: From data collection to structure refinement and beyond Dates : June 18 to 26. Site: Advanced Photon Source, Argonne National Laboratory, Argonne , Illinois (Near Chicago ), USA The school content: Data collection workshop the first two days: beamline training and data collection on GM/CA-CAT beamlines 23ID-B and 23ID-D. For data collection, only the participants' crystals will be used. Software workshop: The rest of the time after data collection will feature many modern crystallographic software packages taught by authors and other experts. It will be organized in three Sections – lectures, tutorials and hands-on trouble-shooting. There will be model data sets available for tutorials but data, provided by participants, will have higher priority for the hands-on sessions. Applicants : Graduate students, postdoctoral researchers and young scientists at the assistant professor level are encouraged to apply. Only 20 applicants will be selected for participation. Participants of the workshop are strongly encouraged to bring their own problem data sets or crystals so the problems can be addressed during data collection workshop and/or hands-on sessions. Application : Application deadline is April 5. The application form, the program, contact info and other details can be found at http: http://www.ccp4.ac.uk/schools/APS-2013/index.php Fees: There is no fee for the workshop. The students will be responsible for their transportation and lodging. The workshop organizers will arrange economical lodging at the Argonne Guest House. The workshop will also cover the expenses for all meals and refreshments. Garib, Ronan and Nukri Ruslan Sanishvili (Nukri) Macromolecular Crystallographer GM/CA@APS X-ray Science Division, ANL 9700 S. Cass Ave. Argonne, IL 60439 Tel: (630)252-0665 Fax: (630)252-0667 rsanishv...@anl.gov
Re: [ccp4bb] a challenge
I actually chose 3dko because it is a kinase (with a ligand), and therefore an interesting candidate for a molecular replacement score. I have not set this up yet, but I think if you look for PDB entries that contain the word kinase and try to molecular-replace all of them into the 3dko dataset, what fraction of them will work? I think that fraction would make a good score for a given molecular replacement pipeline. But, if you want to bootstrap S-SAD phasing with a homolog, then I'd say its definitely cheating if you use a homolog close enough to build your way out of the resulting density without any anomalous information at all. Perhaps the fairest way to do this would be to make a 2-dimensional score? The frac of the dataset you used, plus the BLAST2 E-value of the model you started with vs the 3dko sequence? -James Holton MAD Scientist On Mon, Jan 14, 2013 at 2:31 PM, Nat Echols nathaniel.ech...@gmail.com wrote: On Mon, Jan 14, 2013 at 11:18 AM, Tim Gruene t...@shelx.uni-ac.gwdg.de wrote: I admit not having read all contributions to this thread. I understand the John Henry Challenge as whether there is an 'automated way of producing a model from impossible.mtz'. From looking at it and without having gone all the way to a PDB-file my feeling is one could without too much effort from the baton mode in e.g. coot. This should be even more possible if one also uses existing knowledge about the expected structure of the protein: a kinase domain is quite distinctive. So, James, how much external information from homologous structures are we allowed to use? Running Phaser would certainly be cheating, but if I take (for instance) a 25% identical kinase structure, manually align it to the map and/or a partial model, and use that as a guide to manually rebuild the target model, does that meet the terms of the challenge? -Nat
