Re: [ccp4bb] Question about R/Rfree value difference
I do agree with Tim's reasoning in general, but as Pavel also implied by offering the statistics, I would not be worried about the difference, but by the unreasonably high absolute value of Free R for 2.0 A resolution. I do not think that its simple 'over-fitting' and my worry would not be just the very high difference between R/Rfree, but simply the high Rfree. I would more or less bet that if you check your model in the MolProbity server (http://molprobity.biochem.duke.edu/) it will also have appalling geometry scores(*) Here is what I would suspect and what I would do to check what is wrong: -very incomplete or badly built model: validate in Molprobity, and then rebuild (Coot obviously ...). while doing this check the following which just takes computer time, not yours: -twining: try any de-twining tool, I would simply switch on twining refinement in REFMAC and sit back and read the log carefully. -wrong space group: Try the Zanuda server: http://www.ysbl.york.ac.uk/YSBLPrograms/index.jsp regards - Tassos (*)on the other hand I did bet on Germany winning against Spain, so my betting skills are worse than these of a cephalopod mollusk named 'Paul', http://en.wikipedia.org/wiki/Paul_the_Octopus PS When you use TLS the average B will always go down since a lot of the movement is 'absorbed' by the TLS tensors that describe domain movement - what is in the PDB is just the 'residual' B that cannot be explained by domains moves. On Jul 8, 2010, at 8:32, Tim Gruene wrote: Dear Sampath, You are right, the gap between R and Rfree is significant and indicates that your model was overfitted. Without knowing your data or your model, some reasons for overfitting might be: - you used automated placement of water molecules (e.g. through arpwaters or in coot) and never checked the water molecules for chemical reasonability. How many residues are there in your structure and how many water molecules? - there might be a domain that - despite the resolution - does not resolve with your data but you built somethig nevertheless - you build things into your model while using a too low (approx. 1.0sigma) sigma-level for your map. At too low a contour level you can often see what you _want_ to see in my experience, and not what is there - you screwed up the Rfree set and it's not indendent anymore. However, in that case I would rather expect the difference or the ration to be too small rather than too big. - Your data may be twinned. That's just a first set of reasons but there might be something one could only know by looking at your data. Tim On Wed, Jul 07, 2010 at 10:17:14PM -0700, Sampath Natarajan wrote: Dear all, I have a question about the R free value. I refined a structure with 2A resolution. After model building and restraint refinement using Refmac program, the average B factor was around 50 for all atoms. The R/ Rfree were around 22/34. Then used the TLS refinement choosing entire molecule. Then R/Rfree reduced as 20/32. But the average B factor was reduced as 30. The R/Rfree difference is about 12% in final refinement. I feel it is significantly higher. Could any one suggest me to reduce the Rfree value more? or is it good to submit the data in the PDB database with this 12% difference? Thanks for the suggestions. Sincerely, Sampath N -- -- Tim Gruene Institut fuer anorganische Chemie Tammannstr. 4 D-37077 Goettingen GPG Key ID = A46BEE1A P please don't print this e-mail unless you really need to Anastassis (Tassos) Perrakis, Principal Investigator / Staff Member Department of Biochemistry (B8) Netherlands Cancer Institute, Dept. B8, 1066 CX Amsterdam, The Netherlands Tel: +31 20 512 1951 Fax: +31 20 512 1954 Mobile / SMS: +31 6 28 597791
Re: [ccp4bb] Question about R/Rfree value difference
badly built model reminds me of another possible reason: if you have an old version of ccp4 installed (before 6.1??), there is a default weight of 0.3 for refmac between data and restraints. This value is - in my experience - way too high for normal resolution and at the beginning of refinement resulting, as Tassos described, in a terrible geometry of the model. With recent versions of ccp4, the auto-weighting option in refmac is switched on which usually does a good job. Tim P.S.: Because of this version question one might add to the CCP4 netiquette to always state what program was used and which version of it when asking program specific questions. On Thu, Jul 08, 2010 at 10:14:56AM +0200, Anastassis Perrakis wrote: I do agree with Tim's reasoning in general, but as Pavel also implied by offering the statistics, I would not be worried about the difference, but by the unreasonably high absolute value of Free R for 2.0 A resolution. I do not think that its simple 'over-fitting' and my worry would not be just the very high difference between R/Rfree, but simply the high Rfree. I would more or less bet that if you check your model in the MolProbity server (http://molprobity.biochem.duke.edu/) it will also have appalling geometry scores(*) Here is what I would suspect and what I would do to check what is wrong: -very incomplete or badly built model: validate in Molprobity, and then rebuild (Coot obviously ...). while doing this check the following which just takes computer time, not yours: -twining: try any de-twining tool, I would simply switch on twining refinement in REFMAC and sit back and read the log carefully. -wrong space group: Try the Zanuda server: http://www.ysbl.york.ac.uk/YSBLPrograms/index.jsp regards - Tassos (*)on the other hand I did bet on Germany winning against Spain, so my betting skills are worse than these of a cephalopod mollusk named 'Paul', http://en.wikipedia.org/wiki/Paul_the_Octopus PS When you use TLS the average B will always go down since a lot of the movement is 'absorbed' by the TLS tensors that describe domain movement - what is in the PDB is just the 'residual' B that cannot be explained by domains moves. On Jul 8, 2010, at 8:32, Tim Gruene wrote: Dear Sampath, You are right, the gap between R and Rfree is significant and indicates that your model was overfitted. Without knowing your data or your model, some reasons for overfitting might be: - you used automated placement of water molecules (e.g. through arpwaters or in coot) and never checked the water molecules for chemical reasonability. How many residues are there in your structure and how many water molecules? - there might be a domain that - despite the resolution - does not resolve with your data but you built somethig nevertheless - you build things into your model while using a too low (approx. 1.0sigma) sigma-level for your map. At too low a contour level you can often see what you _want_ to see in my experience, and not what is there - you screwed up the Rfree set and it's not indendent anymore. However, in that case I would rather expect the difference or the ration to be too small rather than too big. - Your data may be twinned. That's just a first set of reasons but there might be something one could only know by looking at your data. Tim On Wed, Jul 07, 2010 at 10:17:14PM -0700, Sampath Natarajan wrote: Dear all, I have a question about the R free value. I refined a structure with 2A resolution. After model building and restraint refinement using Refmac program, the average B factor was around 50 for all atoms. The R/ Rfree were around 22/34. Then used the TLS refinement choosing entire molecule. Then R/Rfree reduced as 20/32. But the average B factor was reduced as 30. The R/Rfree difference is about 12% in final refinement. I feel it is significantly higher. Could any one suggest me to reduce the Rfree value more? or is it good to submit the data in the PDB database with this 12% difference? Thanks for the suggestions. Sincerely, Sampath N -- -- Tim Gruene Institut fuer anorganische Chemie Tammannstr. 4 D-37077 Goettingen GPG Key ID = A46BEE1A P please don't print this e-mail unless you really need to Anastassis (Tassos) Perrakis, Principal Investigator / Staff Member Department of Biochemistry (B8) Netherlands Cancer Institute, Dept. B8, 1066 CX Amsterdam, The Netherlands Tel: +31 20 512 1951 Fax: +31 20 512 1954 Mobile / SMS: +31 6 28 597791 -- -- Tim Gruene Institut fuer anorganische Chemie Tammannstr. 4 D-37077 Goettingen GPG Key ID = A46BEE1A signature.asc Description: Digital signature
Re: [ccp4bb] Question about R/Rfree value difference
Something else to consider is what is your space group ? P212121 but truly P21 with twinning fraction close to 0.5 ? That's one of my recent cases. 1.9 Å data beautifully refined built but the Rwork/Rfree gap was 13 percent. After changing the space group and applying the twin law the gap is 3 percent 18.4 and 21.3 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://web.mac.com/bosch_lab/ On Jul 8, 2010, at 1:17 AM, Sampath Natarajan wrote: Dear all, I have a question about the R free value. I refined a structure with 2A resolution. After model building and restraint refinement using Refmac program, the average B factor was around 50 for all atoms. The R/Rfree were around 22/34. Then used the TLS refinement choosing entire molecule. Then R/Rfree reduced as 20/32. But the average B factor was reduced as 30. The R/Rfree difference is about 12% in final refinement. I feel it is significantly higher. Could any one suggest me to reduce the Rfree value more? or is it good to submit the data in the PDB database with this 12% difference? Thanks for the suggestions. Sincerely, Sampath N
[ccp4bb] idiffdisp - two clicks to define a line does not work on a sector ?
Hi, In idiffdisp (v. 6.1.13 and I presume also in earlier versions), if I read in a bunch of images as a sector, the function 'two clicks to define a line' is not working (it's not lit in fact). This function is quite useful for looking into difficult cases e.g. where weak reflections are ignored by the indexing program. Saving the sector and reading it back in is not an option in idiffdisp either, since it only suggests to save the sector as jpg. In ipdisp, which is no longer supported, this was possible. Is it possible to add them to idiffdisp ? Cheers, Boaz Boaz Shaanan, Ph.D. Dept. of Life Sciences Ben-Gurion University of the Negev Beer-Sheva 84105 Israel Phone: 972-8-647-2220 ; Fax: 646-1710 Skype: boaz.shaanan
Re: [ccp4bb] Calculate the portion of the density within a mask
mapmask will do this. Eleanor See the documentation - it is a bit confalued but certainly works.. Hailiang Zhang wrote: Hi, I want to calculate the portion of the noise density with respect to the whole unit cell (assuming the model is good enough). I plan to first calculate the integral density within the whole unit cell, then build a atom mask around the molecule and calculate the integral density within the mask only. I tried UPPSALA mapman, but it seemed not working as expected. Can somebody refer me some utilities in ccp4 which could accomplish this task? Thanks! Hailiang
Re: [ccp4bb] Question about R/Rfree value difference
Sampath With regard to your question on what sort of statistics you should get within structure determination you might find this service at the PDBe useful : http://www.ebi.ac.uk/pdbe-as/pdbestatistics/PDBeStatistics.jsp You can view and manipulate distributions of R, Rfree and R-Rfree along within many other data distributions from Xray (also NMR/EM) during structure determination. There are also links (clicking the graph) that list all the depositions that have a particular value within the distribution. I agree with Pavel that from your quoted statistics that it would be un-wise to deposit the structure in the current state of refinement as there is clearly an issue. Regards Tom Oldfield Hi Sampath, this is how the distribution of Rwork, Rfree and Rfree-Rwork look like for 'all' PDB structures refined at around 2A resolution. The indicates where your structure stands with respect to this distribution. Histogram of Rwork for models in PDB at resolution 1.90-2.10 A: 0.093 - 0.118 : 2 0.118 - 0.143 : 35 0.143 - 0.168 : 390 0.168 - 0.193 : 1439 0.193 - 0.218 : 1802 your structure 0.218 - 0.242 : 785 0.242 - 0.267 : 159 0.267 - 0.292 : 14 0.292 - 0.317 : 1 0.317 - 0.342 : 1 Histogram of Rfree for models in PDB at resolution 1.90-2.10 A: 0.149 - 0.170 : 10 0.170 - 0.191 : 116 0.191 - 0.213 : 534 0.213 - 0.234 : 1166 0.234 - 0.255 : 1417 0.255 - 0.276 : 942 0.276 - 0.297 : 343 0.297 - 0.319 : 78 0.319 - 0.340 : 17 your structure 0.340 - 0.361 : 5 Histogram of Rfree-Rwork for all model in PDB at resolution 1.90-2.10 A: 0.001 - 0.011 : 41 0.011 - 0.021 : 230 0.021 - 0.031 : 724 0.031 - 0.041 : 1210 0.041 - 0.050 : 1206 0.050 - 0.060 : 654 0.060 - 0.070 : 318 0.070 - 0.080 : 160 0.080 - 0.090 : 56 0.090 - 0.100 : 29 So, it seems your case is the example of typical overfitting, which means the model parameterization or/and the refinement strategy is not good for your data and model. If you send me the data and model files then I will be able (hopefully) to suggest a better refinement strategy or explaine why it's not feasible with available tools. All files will be kept confidentially. The histograms above are obtained using this command from PHENIX family: phenix.r_factor_statistics 2.0 Good luck! Pavel. On 7/7/10 10:17 PM, Sampath Natarajan wrote: Dear all, I have a question about the R free value. I refined a structure with 2A resolution. After model building and restraint refinement using Refmac program, the average B factor was around 50 for all atoms. The R/Rfree were around 22/34. Then used the TLS refinement choosing entire molecule. Then R/Rfree reduced as 20/32. But the average B factor was reduced as 30. The R/Rfree difference is about 12% in final refinement. I feel it is significantly higher. Could any one suggest me to reduce the Rfree value more? or is it good to submit the data in the PDB database with this 12% difference? Thanks for the suggestions. Sincerely, Sampath N
[ccp4bb] Postdoctoral Opportunity at Pfizer Cambridge, MA
There is a postdoctoral position in Structural Biology available at the Pfizer site in Cambridge, Massachusetts. The project involves solving the structures of related signalling protein/receptor complexes and then using computational modeling to design new molecules with enhanced specificity. We are looking for recent Ph.D. recipients with strong skills in biochemistry, x-ray crystallography, and computational crystallography or modeling. Pfizer offers excellent postdoc salary and benefits and has a very open approach to publication. The Cambridge site offers access to the excellent research community in the Boston area. We have a state-of-the art crystallization laboratory and frequent synchrotron access. Interested candidates should send a CV and cover letter to me by email: joel.b...@pfizer.com Best Regards, Joel
Re: [ccp4bb] Twin questions: is my crystals twinned or not?
Hi Partha - A few thoughts: 1. If you attach logs, at least gzip them ... 2. From the fact that Rfree goes down when twinning is switched off, I would think there is no twining. 3. The intensity distribution (moment of E, etc) suggest no twining. The twining operator you use is the crystallographic two fold, I think. 4. During refinement the relative fraction of twin is a refined parameter, to make you model+twining fit the data. The initial twin fraction is only an estimate. 5. Did you consider P3121 or P3221 at all? I think its unlikely, but easy to check in Phaser as alternative space group. 6. Do these R/Rfree include TLS refinement and waters? if not they are quite Ok, given the missing residues 7. I see you only have 430 free reflections. I would argue they are too few to get a reliable Rfree and would aim for more, 10% instead of 5% will give you about 900. Be careful though in not choosing a new Rfree for the refinement from now on. Good luck - A. On 8 Jul 2010, at 20:03, Parthasarathy Sampathkumar wrote: Dear All, Back ground: This is my first experience with a twined dataset. Crystals belong to a small domain of 132 aa, out of which ~40 residues appears to be disordered (~30 of those from C-terminal and C-term His6 tag). Initial space group: P3 with unit cell dimensions: 62.507 62.507 55.117 90.00 90.00 120.00; Resolution 2.35Angs. Pointless suggested P321 (space group # 150) as possibility. I determined structure by MR with Phaser (1 molecule in ASU). After several model building and refinement cycles R and Rfree got stuck at 24.0% and 31.6% respectively. Therefore, I considered P3 space group (now Two molecules in the ASU) with a twin component. I was only able to add handful residues to the model already refined in P321. My current R and Rfree factors are 21.0% and 29.1%, respectively for Two molecules refined in P3 space group. Questions: 1. H-test in cTruncate suggested a twin fraction of 0.43 for the twin operator -h-k, k, -l. Where as Refmac5 with Amplitude based twin refinement gave an initial value of 0.508 for the same operator. Why these values are different between cTruncate and Refmac5 (is this because I asked Refmac5 do amplitude Twin refinement instead of Intensity based)? 2. I noticed in Refmac5 log file that twin fractions changes for every cycle of refinement. During my most recent Refmac5 run Initial estimate of 0.508 for -h-k, k, -l operator changed to 0.504 at the end of 20th cycle. The corresponding values were 0.519 and 0.529, respectively, in a previous round. Since twin estimates were based on measured Intensities (in turn amplitudes) why would they change with refinement (am I missing something here)? 3. When I repeated my final round of Refmac5 WithOut Twin Refinement my R and R-free factors are 22.9% and 28.6%, respectively, which also appears to be OK for 2.35 Angs. data (in fact, slightly better R-free). These values are likely to improve little bit after completing the solvent model. So, is this crystal really twinned? I have attached log files of cTruncate and most recent Refmac5 run with Twin refinement. Apologies for attachments (at least no image files). Thank you all in advance for educating me. -Partha 34_truncate_anl.log40_refmac5.log
Re: [ccp4bb] Twin questions: is my crystals twinned or not?
On Thursday 08 July 2010 11:03:30 am Parthasarathy Sampathkumar wrote: Dear All, Back ground: This is my first experience with a twined dataset. Crystals belong to a small domain of 132 aa, out of which ~40 residues appears to be disordered (~30 of those from C-terminal and C-term His6 tag). Initial space group: P3 with unit cell dimensions: 62.507 62.507 55.117 90.00 90.00 120.00; Resolution 2.35Angs. Pointless suggested P321 (space group # 150) as possibility. I determined structure by MR with Phaser (1 molecule in ASU). After several model building and refinement cycles R and Rfree got stuck at 24.0% and 31.6% respectively. OK. Therefore, I considered P3 space group (now Two molecules in the ASU) with a twin component. But now you veer off track. P3 + 0.5 twinning fraction is the same as P321, except that you have doubled the number of refined parameters by modelling two copies of the protein rather than one. This is probably not what you want to do. I was only able to add handful residues to the model already refined in P321. My current R and Rfree factors are 21.0% and 29.1%, respectively for Two molecules refined in P3 space group. Turning on twin refinement in refmac is always expected to drop R slightly, regardless of whether the structure is actually twinned or not. I'll leave it to Garib to explain why this is the case :-) In other words, given that you doubled the number of parameters and also turned on twin refinement, a drop in Rfree from .31 to .29 is not impressive. I don't think that any of the information you present justifies treating the structure as being in P3. Instead you should be considering the possibility that the structure really is in P321, but there is a twin law that makes it appear partially hexagonal. See the table of possible twin laws for P3(?)21: http://www.ccp4.ac.uk/html/twinning.html Questions: 1. H-test in cTruncate suggested a twin fraction of 0.43 for the twin operator -h-k, k, -l. Was this relative to a P3 indexing, or a P321 indexing? Where as Refmac5 with Amplitude based twin refinement gave an initial value of 0.508 for the same operator. That's strange. A twinning fraction should never be higher than 0.5 But let's get your true twinning law sorted out first before worrying about whether refmac should report this as 0.508 or 0.492 Ethan Why these values are different between cTruncate and Refmac5 (is this because I asked Refmac5 do amplitude Twin refinement instead of Intensity based)? 2. I noticed in Refmac5 log file that twin fractions changes for every cycle of refinement. During my most recent Refmac5 run Initial estimate of 0.508 for -h-k, k, -l operator changed to 0.504 at the end of 20th cycle. The corresponding values were 0.519 and 0.529, respectively, in a previous round. Since twin estimates were based on measured Intensities (in turn amplitudes) why would they change with refinement (am I missing something here)? 3. When I repeated my final round of Refmac5 WithOut Twin Refinement my R and R-free factors are 22.9% and 28.6%, respectively, which also appears to be OK for 2.35 Angs. data (in fact, slightly better R-free). These values are likely to improve little bit after completing the solvent model. So, is this crystal really twinned? I have attached log files of cTruncate and most recent Refmac5 run with Twin refinement. Apologies for attachments (at least no image files). Thank you all in advance for educating me. -Partha -- Ethan A Merritt Biomolecular Structure Center, K-428 Health Sciences Bldg University of Washington, Seattle 98195-7742
[ccp4bb] Beginning crystallography text
Hi all, I haven't gotten past the phase of growing the crystal, but I'd certainly still like to learn the actual theories of crystallography. Can anyone recommend a good beginner to mid-level text on macromolecular crystallography? Thanks, Peter
Re: [ccp4bb] Beginning crystallography text
This book worked great for me: http://www.amazon.com/Crystallography-Made-Crystal-Clear-Third/dp/0125870736/ref=sr_1_3?ie=UTF8s=booksqid=1278618216sr=1-3 Cheers, Thomas On Thu, Jul 8, 2010 at 12:35, Peter Hsu hsuu...@u.washington.edu wrote: Hi all, I haven't gotten past the phase of growing the crystal, but I'd certainly still like to learn the actual theories of crystallography. Can anyone recommend a good beginner to mid-level text on macromolecular crystallography? Thanks, Peter
Re: [ccp4bb] Beginning crystallography text
At the risk of appearing immodest: http://www.amazon.com/Protein-Crystallography-Eaton-E-Lattman/dp/0801888069/ref=sr_1_10?ie=UTF8s=booksqid=1278618335sr=1-10 On 8 Jul 2010, at 3:35 PM, Peter Hsu wrote: Hi all, I haven't gotten past the phase of growing the crystal, but I'd certainly still like to learn the actual theories of crystallography. Can anyone recommend a good beginner to mid-level text on macromolecular crystallography? Thanks, Peter --- Patrick J. Loll, Ph. D. Professor of Biochemistry Molecular Biology Director, Biochemistry Graduate Program Drexel University College of Medicine Room 10-102 New College Building 245 N. 15th St., Mailstop 497 Philadelphia, PA 19102-1192 USA (215) 762-7706 pat.l...@drexelmed.edu
[ccp4bb] RES: [ccp4bb] Beginning crystallography text
Hi Peter, I understood your situation and I believe the best literature for you is the Dr. Rupp's Book: Biomolecular Crystallography: Principles, Practice, and Application to Structural Biology Author: Benhard Rupp. Good luck in your studies. Best regards, Júlio César -Mensagem original- De: CCP4 bulletin board [mailto:ccp...@jiscmail.ac.uk] Em nome de Peter Hsu Enviada em: quinta-feira, 8 de julho de 2010 16:36 Para: CCP4BB@JISCMAIL.AC.UK Assunto: [ccp4bb] Beginning crystallography text Hi all, I haven't gotten past the phase of growing the crystal, but I'd certainly still like to learn the actual theories of crystallography. Can anyone recommend a good beginner to mid-level text on macromolecular crystallography? Thanks, Peter
Re: [ccp4bb] Beginning crystallography text
Having recently completed the CSHL Macromolecular crystallography course, I can recommend Introduction to Macromolecular Crystallography by Alexander McPherson (ISBN 987-0-470-18590-2). I am posting the link below: http://www.amazon.com/Introduction-Macromolecular-Crystallography-Alexander-McPherson/dp/0470185902/ref=sr_1_1?ie=UTF8s=booksqid=1278619717sr=1-1 Kind regards and good luck! Bryan -- Confidentiality Notice: This message is private and may contain confidential and proprietary information. If you have received this message in error, please notify us and remove it from your system and note that you must not copy, distribute or take any action in reliance on it. Any unauthorized use or disclosure of the contents of this message is not permitted and may be unlawful. -Original Message- From: CCP4 bulletin board [mailto:ccp...@jiscmail.ac.uk] On Behalf Of Peter Hsu Sent: Thursday, July 08, 2010 3:36 PM To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] Beginning crystallography text Hi all, I haven't gotten past the phase of growing the crystal, but I'd certainly still like to learn the actual theories of crystallography. Can anyone recommend a good beginner to mid-level text on macromolecular crystallography? Thanks, Peter
Re: [ccp4bb] Beginning crystallography text
I like David Blow's book for beginners -- one can get the gist of things without having much math: http://www.abebooks.com/servlet/SearchResults?an=blowsts=ttn=crystallographyx=35y=6 Bernhard Rupp's book, mentioned earlier, is the current gold standard, in my view. Bob On Thu, 8 Jul 2010, Prince, D Bryan wrote: Having recently completed the CSHL Macromolecular crystallography course, I can recommend Introduction to Macromolecular Crystallography by Alexander McPherson (ISBN 987-0-470-18590-2). I am posting the link below: http://www.amazon.com/Introduction-Macromolecular-Crystallography-Alexander-McPherson/dp/0470185902/ref=sr_1_1?ie=UTF8s=booksqid=1278619717sr=1-1 Kind regards and good luck! Bryan -- Confidentiality Notice: This message is private and may contain confidential and proprietary information. If you have received this message in error, please notify us and remove it from your system and note that you must not copy, distribute or take any action in reliance on it. Any unauthorized use or disclosure of the contents of this message is not permitted and may be unlawful. -Original Message- From: CCP4 bulletin board [mailto:ccp...@jiscmail.ac.uk] On Behalf Of Peter Hsu Sent: Thursday, July 08, 2010 3:36 PM To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] Beginning crystallography text Hi all, I haven't gotten past the phase of growing the crystal, but I'd certainly still like to learn the actual theories of crystallography. Can anyone recommend a good beginner to mid-level text on macromolecular crystallography? Thanks, Peter -- = Robert M. Sweet E-Dress: sw...@bnl.gov Group Leader, PXRR: Macromolecular ^ (that's L Crystallography Research Resource at NSLSnot 1) http://px.nsls.bnl.gov/ Biology Dept Brookhaven Nat'l Lab. Phones: Upton, NY 11973631 344 3401 (Office) U.S.A. 631 344 2741 (Facsimile) =
[ccp4bb] Computational Crystallography Newsletter - Issue 1
Dear Colleagues, I am pleased to announce the publication of the first issue of the Computational Crystallography Newsletter: http://www.phenix-online.org/newsletter/ It features articles, meeting announcements and reports, information on research and other items of interest to crystallographers and users of crystallographic software. The target audience is structural biologists including both students and experts. Although this first issue has many Phenix and cctbx related articles we would like this to be a resource for the entire community and expect to see a broad range of topics covered in future issues. Anyone interested in publishing articles or other items in the newsletter is encouraged to contact the editor at any time. Submission of text by email using the word-processing templates available online is requested. Cheers, Paul -- Paul Adams Deputy Division Director, Physical Biosciences Division, Lawrence Berkeley Lab Adjunct Professor, Department of Bioengineering, U.C. Berkeley Vice President for Technology, the Joint BioEnergy Institute Head, Berkeley Center for Structural Biology Building 64, Room 248 Tel: 1-510-486-4225, Fax: 1-510-486-5909 http://cci.lbl.gov/paul Lawrence Berkeley Laboratory 1 Cyclotron Road BLDG 64R0121 Berkeley, CA 94720, USA. Executive Assistant: Marian Harris [ mshar...@lbl.gov ] [ 1-510-486-6886 ] --
[ccp4bb] How to make fft-map more physically meaningful?
Hi there: I found that the grid values in the map file generated by CCP4-fft generally has a mean value of ~0, and of course there will be lots of negative values. This apparently is not the real physics, since the electron density has to be positive everywhere (hope I am right). Can somebody give me any hint how to convert the fft map file which has mean value of 0, to a more physically meaningful map which has positive densities everywhere? (I thought about offsetting the whole map by the minimum negative values to make everything positive, but I doubt it is right). Best Regards, Hailiang
Re: [ccp4bb] Beginning crystallography text
Back when I was a graduate student, my favorite book was Drenth. However, that book was never a favorite with most students, who preferred Crystallography Made Crystal Clear. I also think the Blow book is good. I'm not familiar with the newer books written by our mailing list colleagues. ho
Re: [ccp4bb] How to make fft-map more physically meaningful?
Hailiang Zhang wrote: Hi there: I found that the grid values in the map file generated by CCP4-fft generally has a mean value of ~0, and of course there will be lots of negative values. This apparently is not the real physics, since the electron density has to be positive everywhere (hope I am right). Can somebody give me any hint how to convert the fft map file which has mean value of 0, to a more physically meaningful map which has positive densities everywhere? (I thought about offsetting the whole map by the minimum negative values to make everything positive, but I doubt it is right). Best Regards, Hailiang Actually taking the minimum value as zero might be a good approximation, as long as the resolution is high enough so there are gaps in the protein too small to be solvent-filled but large enough to be resolved from surrounding density. Maps from FFT will always have average value zero unless you include the 0,0,0 reflection: the transform is a sum of sin and cos terms, all of which have zero value when integrated over the unit cell, except the cos(0.X) term. So any linear combination of these terms will average to zero if it doesn't include the zero order term. The 0,0,0 reflection is hard or impossible to measure because it gets mixed up with the undiffracted beam. But it is easy to calculate, because the integral of unity against the electron density is just the average electron density times the volume, or the total number of electrons. So if you know the total number of electrons in the unit cell, you can divide by the unit cell volume to get the average electron density (OK, I guess that is obvious) and add it to the zero-average FFT map. This assumes the map is on an absolute scale, which won't be quite true, so your idea of offsetting the minimum to zero may be more satisfactory. Ed
Re: [ccp4bb] How to make fft-map more physically meaningful?
Edward A. Berry wrote: Hailiang Zhang wrote: Hi there: I found that the grid values in the map file generated by CCP4-fft generally has a mean value of ~0, and of course there will be lots of negative values. This apparently is not the real physics, since the electron density has to be positive everywhere (hope I am right). Can somebody give me any hint how to convert the fft map file which has mean value of 0, to a more physically meaningful map which has positive densities everywhere? (I thought about offsetting the whole map by the minimum negative values to make everything positive, but I doubt it is right). Best Regards, Hailiang Actually taking the minimum value as zero might be a good approximation, as long as the resolution is high enough so there are gaps in the protein too small to be solvent-filled but large enough to be resolved from surrounding density. Maps from FFT will always have average value zero unless you include the 0,0,0 reflection: the transform is a sum of sin and cos terms, all of which have zero value when integrated over the unit cell, except the cos(0.X) term. So any linear combination of these terms will average to zero if it doesn't include the zero order term. The 0,0,0 reflection is hard or impossible to measure because it gets mixed up with the undiffracted beam. But it is easy to calculate, because the integral of unity against the electron density is just the average electron density times the volume, or the total number of electrons. So if you know the total number of electrons in the unit cell, you can divide by the unit cell volume to get the average electron density (OK, I guess that is obvious) and add it to the zero-average FFT map. This assumes the map is on an absolute scale, which won't be quite true, so your idea of offsetting the minimum to zero may be more satisfactory. Ed Ed is right, of course. Just remember to include ALL the electrons in the unit cell - both those of the protein and those of the solvent, ordered and disordered. Dale Tronrud
Re: [ccp4bb] Beginning crystallography text
The question of what textbook to use is very much context sensitive, that is, it depends on what the reader wants and needs to know. Unfortunately, this question us easy to answer with hindsight, but not so obvious to the person looking for answers. Having said that, I declare a conflict of interest as one of the mentioned textbook authors. The conflict, however, is modest because I am not aware of anyone making a fortune on crystallography textbooks. I think it is reasonable to delineate the textbook market by what the reader ultimately wants to accomplish. What a structural biologist should know, versus what is expert knowledge, has been a contentious issue for quite some years. A lot of people have thought hard about this, and the education committees of the American Crystallographic Association (ACA) and USNC/Cr organized a crystallography education summit, whose outcome is the consensus policy statement on crystallography education and training available from here: http://www.ruppweb.org/garland/study_group.htm I privately think that as a first contact for the user of structure models, the Rhodes book is a great start. If you are a tad more interested in how it works, Jenny Glusker's old text in its revised form is still one of my favorites, and the Blow book as well Alex's compilation are quite useful. Drenth helps once you are already engaged in the business, and have some idea what it is about. The IUCr compilation is an extremely useful hard core resource if you are interested in the nuz and bolz of crystallography in general. Not to forget the excellent multi-author volumes of Methods of Enzymology as an in-depth resource. Having said that, the reason why I decided to add another tome (BMC) to the already prolific writings in protein crystallography is that I felt that none of the above provided a consistent and modern picture of crystallography in the probabilistic framework it actually operates in. This is - in a crystallographic time frame - ancient history; a first resource being the 1952 work of Crick and Blow, and it continues via French and Wilson to Bricogne and on. So, as a concluding statement, I think there is more to biomolecular crystallography that just nuz and bolz, and it touches many very fundamental challenges and uncertainties, ultimately forcing my emphasis on probabilistic approaches and the resulting digressions in the subversive sidebars. Consequentially, if you like the Schaum series and Kaplan SAT rest prep books, don't waste your money on my book. Instead, get one of the (nearly as expensive by weight and volume) monographs mentioned above, they are in fact good and will lead you in the right direction. If you like Neal Stevenson, Rev. Bayes, and a touch of randomness, and you understand that the probability of receiving the Nobel Price approaches practically zero once you have been infected by the spirit of BMC, go for it ;-) BR -Original Message- From: CCP4 bulletin board [mailto:ccp...@jiscmail.ac.uk] On Behalf Of Robert Sweet Sent: Thursday, July 08, 2010 1:26 PM To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] Beginning crystallography text I like David Blow's book for beginners -- one can get the gist of things without having much math: http://www.abebooks.com/servlet/SearchResults?an=blowsts=ttn=crystallograp hyx=35y=6 Bernhard Rupp's book, mentioned earlier, is the current gold standard, in my view. Bob On Thu, 8 Jul 2010, Prince, D Bryan wrote: Having recently completed the CSHL Macromolecular crystallography course, I can recommend Introduction to Macromolecular Crystallography by Alexander McPherson (ISBN 987-0-470-18590-2). I am posting the link below: http://www.amazon.com/Introduction-Macromolecular-Crystallography-Alexander- McPherson/dp/0470185902/ref=sr_1_1?ie=UTF8s=booksqid=1278619717sr=1-1 Kind regards and good luck! Bryan -- Confidentiality Notice: This message is private and may contain confidential and proprietary information. If you have received this message in error, please notify us and remove it from your system and note that you must not copy, distribute or take any action in reliance on it. Any unauthorized use or disclosure of the contents of this message is not permitted and may be unlawful. -Original Message- From: CCP4 bulletin board [mailto:ccp...@jiscmail.ac.uk] On Behalf Of Peter Hsu Sent: Thursday, July 08, 2010 3:36 PM To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] Beginning crystallography text Hi all, I haven't gotten past the phase of growing the crystal, but I'd certainly still like to learn the actual theories of crystallography. Can anyone recommend a good beginner to mid-level text on macromolecular crystallography? Thanks, Peter -- = Robert M. Sweet E-Dress:
