Re: [ccp4bb] Charge flipping.
Dear Francis, I am very interested in your work with this. In the paper :- A. Mukherjee, J.R. Helliwell and P. Main 'The use of MULTAN to locate the positions of anomalous scatterers'. (1989) Acta Cryst. *A45*, 715-718. you will see that missing centric reflections was not limiting. However, the more dilute the anomalous scatterers versus the protein atoms led to the practical consequence of increasing the number of required iterations to a successful answer. That said the potential effect of missing centric data and the influence of random and systematic errors on the charge flipping may of course be different. Best wishes, John Prof John R helliwell On Sat, May 22, 2010 at 3:23 PM, Francis E Reyes francis.re...@colorado.edu wrote: Hi all I've been playing around with charge flipping for macromolecular substructure determination with pretty promising results. I'm particularly attracted to the fact that it solves structures in P1, with no space group assumptions and curious how it would handle some of the pseudosymmetry cases I've come into in my time. I'd like to know if anyone's had experience with this method, and open up the discussion with the following questions: As the algorithm starts with completely random phases and charge flips the map in P1, what is the importance of measuring (good or any) anomalous signal at all (for the sole purpose of finding the heavy atoms)? At first pass it would seem that just as long as you have an incorporated heavy atom and the density of that region is greater than delta, that this alone would be sufficient for locating the position of the heavy atom. In other words just as long as your heavy atom is sufficiently higher in contrast than your protein/rna it would be a good enough criteria. In the above regime, would the importance of measuring anomalous data be more important for substructure refinement (via phaser, mlphare, sharp, solve/resolve)? Now to a more specific question for those who've had experience (or maybe the authors are subscribed here): Orthorhombic C2221 using SUPERFLIP heavy atoms are found with great peakiness (before noise suppression: peakiness = 5, after noise suppression peakiness 25, good separation of heavy atom peaks from noise peaks in resulting pdb). Yet the space group check via the sym operators is rather poor (overall agreement close to 100). My interpretation is that the heavy atoms are found, but the space group is wrong? Thanks! F - Francis Reyes M.Sc. 215 UCB University of Colorado at Boulder gpg --keyserver pgp.mit.edu --recv-keys 67BA8D5D 8AE2 F2F4 90F7 9640 28BC 686F 78FD 6669 67BA 8D5D -- Professor John R Helliwell DSc
Re: [ccp4bb] Can CCP4 print the sigma level of the electron density at each residue or atom center?
If you ask for Real space R factor from overlapmap it gives you the mean density for eaxch residue main chain and side chain, or mean density at atom centre. It is rather confusedly labelled Fobs(mc) etc. You could convert this to sigma level by just dividing the values. Eleanor The Hailiang Zhang wrote: Hi, Phenix.model_vs_data offers a great function which prints out the sigma level of the electron density at each residue or atom center. This is very useful comparing the relative density between two maps at the given region, however, it is running a little bit slow. I am just wondering whether some CCP4 subroutines also provides the similar function (didn't find in overlapmap). Thanks a lot! Best Regards, Hailiang
Re: [ccp4bb] Finding best model for molecular replacement
Some afterthoughts: Of course avoid the common MR problems - assigning wrong SG, saturating low resolution data etc etc.. 1) Any sequence search tool might have told you there was only a poor match available. 23% is very marginal for MR and with that degree of similarity you are very wise to start searching for exptl phase methods. However it is sometimes possible to get an MR solution which can at least be helpful as a guide while you build your model into the dexptlly phased map! BALBES does a very good job at analysing likely models - could your sequence suggestr a multimer? are there domains which fit well? etc etc. Then there are the other methods which can give answers - aligning multiple models, doing homology modelling etc. But even if you found a reasonable solution rebuilding and refinement can be difficult - much easier to have some exptl phases to guide that.. Eleanor Ashley Buckle wrote: This is generally a good idea, but removal of residues is a subjective process, and a little trial and error. If your sequence searching finds multiple search models you can superimpose them and systematically remove the poorer fitting regions based upon RMSD. We have built a server for this at http://pxgrid.med.monash.edu.au:8080/mustangserver/ Also see the paper: http://www.plosone.org/article/info%3Adoi%2F10.1371%2Fjournal.pone.0010048 This can generate A LOT of search models, so you need a way of testing them all, and possibly varying other parameters (eg in PHASER varying RMSD, space groups to test, clashes). This can generate 100 runs, so best to be able to use a compute cluster, and some way of queing the jobs (eg MrBump, BALBES etc. Another solution is here: http://www.plosone.org/article/info%3Adoi%2F10.1371%2Fjournal.pone.0010049 hope that helps Ashley On 25/05/2010, at 2:52 AM, Jürgen Bosch wrote: You've also applied BRAIN 2.0 ? I mean looked at homologous structures, superimposed them and decided which parts are to be removed ? Never trust programs :-) There could be a flexible alpha helix which if you removed it would have given you in all programs a solution. it's Monday, Jürgen On May 24, 2010, at 10:24 AM, Paul Lindblom wrote: The last molrep job just finished and it found only an odd solution. So I think I will try to get my phases elsewhere. But I am somewhat astonished that there are still enough cases you can't solve by MR. Thanks to all who replied. Here is a list of servers/programs to find a MR model: http://www.ebi.ac.uk/pdbe-srv/view/ http://www.ebi.ac.uk/Tools/fasta33/index.html http://meta.bioinfo.pl/submit_wizard.pl XtalPred http://ffas.burnham.org/XtalPred-cgi/xtal.pl Balbes http://www.ysbl.york.ac.uk/~fei/balbes/ use the OCA browser for FASTA searches of the PDB Modeller or Rosetta (both also available as web servers) ensemble of many proteins with Phaser FFAS server maintained by the Godzik lab generate some models using the Phyre server ( http://www.sbg.bio.ic.ac.uk/~phyre/ ) and feed the best .pdbs into Mr Bump. - 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/ Associate Professor Ashley M Buckle NHMRC Senior Research Fellow The Department of Biochemistry and Molecular Biology, Faculty of Medicine Monash University, Clayton, Vic 3800 Australia http://www.med.monash.edu.au/biochem/staff/abuckle.html iChat/AIM: blindcaptaincat skype: ashley.buckle Tel: (613) 9902 9313 (office) Fax : (613) 9905 4699 mobile: 0430 913031
Re: [ccp4bb] Alignment software
Hy Mohd, There was recently a nice review about all the alignment software available. You might find one you like in there. Visualization of multiple alignments, phylogenies and gene family evolution. http://www.nature.com/nmeth/journal/v7/n3s/abs/nmeth.1434.html By the way there's also a review on structures visualization software in the same supplement issue about visualizing biological data, that might interest other readers form that list as well. Visualization of macromolecular structures. http://www.nature.com/nmeth/journal/v7/n3s/abs/nmeth.1427.html Best, Luc -- Luc Bonnefond, PhD. Division of Structure Biology Department of Basic Medical Sciences, Institute of Medical Science, The University of Tokyo bonne...@ims.u-tokyo.ac.jp l.bonnef...@gmail.com
Re: [ccp4bb] Fwd: calculate the real space R factor using OVERLAPMAP
On Tue, May 25, 2010 at 6:22 AM, Frank von Delft frank.vonde...@sgc.ox.ac.uk wrote: Hi Ian, I read with great interest. But got stumped here: - how you compute sigma(rho)? See my reply to George Sheldrick's post. I think your reply did not make it out to the BB, certainly neither to my inbox nor to the archive. Do you think you could post it again? Hi Frank, my apologies, I failed to spot that my reply wasn't being copied to the BB, here it is: I just use the EXTENDS program (I modified Phil Evans' original EXTEND program), which gets an initial estimate as the RMS difference density of the map from FFT (i.e. either an a.u. or a complete cell) as you say, then makes a correction for outliers ( 3 sigma), replacing the RMSD value in the map header with the estimated standard uncertainty (i.e. sigma). This is not perfect in terms of outlier rejection: to do it rigorously takes significantly more CPU time (i.e. more than a few seconds), and I wanted to keep map calculation fast (since we do a lot of it!), but it's probably good enough. By the way, I'm afraid you're wrong with this statement: Note that I'm not proposing anything new, this is all explained in standard statistics textbooks (Kendall's Advanced Theory of Statistics by Stuart Ord is probably the best). In fact this is exactly my point: why re-invent the wheel (and likely end up with a square one!) when the appropriate statistics is all there in the textbooks and has been for ~ 80 years? Any time someone spots that an old technique is relevant in a new context, that is something new. And since you bemoan that this is not being used, that by definition makes it a new context :) I'm happy to go along with that! So I'll be bugging Paul to put this in coot, e.g. for sidechain fitting. I think he already got the message! Cheers -- Ian
Re: [ccp4bb] Alignment software
Thank you all for very helpful suggestions, I used the software STRAP, Espript is also user friendly and effective. Many recommended ALINE, but unfortunately I had troubles with the download. Thanks again, Mohd
Re: [ccp4bb] Finding best model for molecular replacement
Dear Paul, A few options may be of interest: RCSB PDB Sequence Searching: Paste your sequence into the sequence search box at: http://www.pdb.org/pdb/search/advSearch.do?st=SequenceQuery (also available from Advanced Search and from the Sequence Search of the left hand menu). Sequence search results are ordered by e-value. Note, the sequence alignment on the results page comes from the BLAST output file. BLAST uses the sequence from SEQRES and the sequence numbering always starts at 1. The sequence numbers of the residues in the ATOM records may be different. To see which part of an aligned sequence is actually present in a structure, go to the Structure Summary page for an entry and click on the Sequence tab. PSI Structural Genomics Knowledgebase: Paste your sequence into the search box at http://kb.psi-structuralgenomics.org/ This search will return any related structures, models, targets, protocols, and materials. MRBUMP (CCP4: Supported Program): http://www.ccp4.ac.uk/MrBUMP/mrbump_doc.html For a given target sequence, automated discovery of chains, domains and multimers that are possible templates for molecular replacement search models Christine RCSB Protein Data Bank On 05/23/10 15:23, Paul Lindblom wrote: Hi everybody, I just crystallized a new project protein. How can I find a possible model for using molecular replacement? I have the sequence of my protein. Is it enough to make a sequence search in the pdb? Or is there another approach I can use? Thanks a lot, Paul
Re: [ccp4bb] Alignment software
Hi there, I remember a friend of mine showing me Aline, and it looked very nice. However, I can't make it work on my MacBook Pro (Mac OS X 10.6.3). With respect to the instruction I found on the website, I had to change the export lines to be added to the .bashrc in order not to get error messages. That's how I added the path info: export ALINEHOME='/sw64/share/xtal/aline_011208' export PATH=$PATH:$ALINEHOME/bin however, I still get an error while trying to run aline: s...@host005:~ aline Can't locate Tk.pm in @INC (@INC contains: /sw64/lib/perl5 /sw64/lib/perl5/darwin /Library/Perl/Updates/5.10.0 /System/Library/Perl/5.10.0/darwin-thread-multi-2level /System/Library/Perl/5.10.0 /Library/Perl/5.10.0/darwin-thread-multi-2level /Library/Perl/5.10.0 /Network/Library/Perl/5.10.0/darwin-thread-multi-2level /Network/Library/Perl/5.10.0 /Network/Library/Perl /System/Library/Perl/Extras/5.10.0/darwin-thread-multi-2level /System/Library/Perl/Extras/5.10.0 .) at /sw64/share/xtal/aline_011208/bin/aline line 37. BEGIN failed--compilation aborted at /sw64/share/xtal/aline_011208/bin/aline line 37. s...@host005:~ I even tried with the perl -MCPAN -e 'install Tk' command, as suggested by the website, but nothing changed. Any idea of what's wrong here? Thanks a lot for the tips, ciao s On May 25, 2010, at 5:47 AM, Charlie Bond wrote: Hi Victor, Thanks for recommending ALINE. It is not exactly true that development has stopped. It is just glacially slow as funding for such utilities is not easily obtained! Recommendations for improvements (to improve that potential) are well-received and a keen user can write their own plug-ins to achieve new functionality. Cheers, Charlie Victor Alves wrote: Hi Mohd You can also check: ALINE (An Extensible WYSIWYG Protein Sequence Alignment Editor for Publication Quality Figures) http://crystal.bcs.uwa.edu.au/px/charlie/software/aline/ It’s quite nice and has a lot of potential, although unfortunately it seems that development of the software as stopped :-( Cheers Victor Alves *From:* CCP4 bulletin board [mailto:ccp...@jiscmail.ac.uk] *On Behalf Of *Salameh, Mohd A., Ph.D. *Sent:* sexta-feira, 21 de Maio de 2010 16:11 *To:* CCP4BB@JISCMAIL.AC.UK *Subject:* [ccp4bb] Alignment software Dear All, I’m trying to prepare an alignment figure of 2 proteins that highlight conserved and similar residues and probably secondary structures; I will greatly appreciate it if anybody can recommend a software that I can use. Thanks, Mohd -- Charlie Bond Professorial Fellow University of Western Australia School of Biomedical, Biomolecular and Chemical Sciences M310 35 Stirling Highway Crawley WA 6009 Australia charles.b...@uwa.edu.au +61 8 6488 4406 -- Sebastiano Pasqualato, PhD IFOM-IEO Campus Dipartimento di Oncologia Sperimentale Istituto Europeo di Oncologia via Adamello, 16 20139 - Milano Italy tel +39 02 9437 5094 fax +39 02 9437 5990
Re: [ccp4bb] UPPSALA: ERROR --- Serious FRCSYM error
Hailiang Zhang wrote: Hi, When I run UPPSALA rsfit, there are lots of ERROR --- Serious FRCSYM error. These atoms/residues are generally around the protein surface, so I guess the reason is the mask were out of the unit cell. Is there any way to avoid this? I've seen that error in averaging/expanding. As you suggest it seems to be the voxel is on the edge, and one more layer beyond the exact asymmetric unit needs to be added. In my case it could be fixed with ccp4 mapmask, something like (for GRID 280 340 456) #2.MAPMASK: mapmask MAPIN temp.map MAPOUT chg1.map END-extend xyzlim 0 280 0 340 0 114 END-extend here this is exactly one asymmetric unit (P212121)- If you still get fracsym's add one layer to each dimension until it behaves? Ed
Re: [ccp4bb] Alignment software
Hi there, I just got this working a couple of days ago under Mac OS X 10.6.3. I manually compiled and installed Perl-Tk from CPAN. Steps taken: 1. Download and ungzip / untar: http://search.cpan.org/CPAN/authors/id/S/SR/SREZIC/Tk-804.028_501.tar.gz(the newer -503 version didn't work) 2. Change into the directory 3. Execute perl Makefile.PL 4. make 5. make test 6. sudo make install Hopefully that should get things working. Cheers, Roger On 25 May 2010 17:50, Sebastiano Pasqualato sebastiano.pasqual...@ifom-ieo-campus.it wrote: Hi there, I remember a friend of mine showing me Aline, and it looked very nice. However, I can't make it work on my MacBook Pro (Mac OS X 10.6.3). With respect to the instruction I found on the website, I had to change the export lines to be added to the .bashrc in order not to get error messages. That's how I added the path info: export ALINEHOME='/sw64/share/xtal/aline_011208' export PATH=$PATH:$ALINEHOME/bin however, I still get an error while trying to run aline: s...@host005:~ aline Can't locate Tk.pm in @INC (@INC contains: /sw64/lib/perl5 /sw64/lib/perl5/darwin /Library/Perl/Updates/5.10.0 /System/Library/Perl/5.10.0/darwin-thread-multi-2level /System/Library/Perl/5.10.0 /Library/Perl/5.10.0/darwin-thread-multi-2level /Library/Perl/5.10.0 /Network/Library/Perl/5.10.0/darwin-thread-multi-2level /Network/Library/Perl/5.10.0 /Network/Library/Perl /System/Library/Perl/Extras/5.10.0/darwin-thread-multi-2level /System/Library/Perl/Extras/5.10.0 .) at /sw64/share/xtal/aline_011208/bin/aline line 37. BEGIN failed--compilation aborted at /sw64/share/xtal/aline_011208/bin/aline line 37. s...@host005:~ I even tried with the perl -MCPAN -e 'install Tk' command, as suggested by the website, but nothing changed. Any idea of what's wrong here? Thanks a lot for the tips, ciao s On May 25, 2010, at 5:47 AM, Charlie Bond wrote: Hi Victor, Thanks for recommending ALINE. It is not exactly true that development has stopped. It is just glacially slow as funding for such utilities is not easily obtained! Recommendations for improvements (to improve that potential) are well-received and a keen user can write their own plug-ins to achieve new functionality. Cheers, Charlie Victor Alves wrote: Hi Mohd You can also check: ALINE (An Extensible WYSIWYG Protein Sequence Alignment Editor for Publication Quality Figures) http://crystal.bcs.uwa.edu.au/px/charlie/software/aline/ It’s quite nice and has a lot of potential, although unfortunately it seems that development of the software as stopped :-( Cheers Victor Alves *From:* CCP4 bulletin board [mailto:ccp...@jiscmail.ac.uk] *On Behalf Of *Salameh, Mohd A., Ph.D. *Sent:* sexta-feira, 21 de Maio de 2010 16:11 *To:* CCP4BB@JISCMAIL.AC.UK *Subject:* [ccp4bb] Alignment software Dear All, I’m trying to prepare an alignment figure of 2 proteins that highlight conserved and similar residues and probably secondary structures; I will greatly appreciate it if anybody can recommend a software that I can use. Thanks, Mohd -- Charlie Bond Professorial Fellow University of Western Australia School of Biomedical, Biomolecular and Chemical Sciences M310 35 Stirling Highway Crawley WA 6009 Australia charles.b...@uwa.edu.au +61 8 6488 4406 -- Sebastiano Pasqualato, PhD IFOM-IEO Campus Dipartimento di Oncologia Sperimentale Istituto Europeo di Oncologia via Adamello, 16 20139 - Milano Italy tel +39 02 9437 5094 fax +39 02 9437 5990 -- Roger B. Dodd, PhD. Cambridge Institute for Medical Research Wellcome Trust/MRC Building Hills Road Cambridge CB2 0XY UK
[ccp4bb] Structure Based Sequence Alignment
Dear All; Can some body tell me a website for structure based sequence alignment, which can also pin point the similar and identical residues in different colors. regards Bashir -- Muhammad Bashir Khan Department for Biomolecular Structural Chemistry Max F. Perutz Laboratories University of Vienna Campus Vienna Biocenter 5 A-1030 Vienna Austria Phone: +43(1)427752224 Fax: +43(1)42779522
[ccp4bb] how to read the ascii map converted by MAPTONA4 and MAPEXCHANGE
Hi, I wanted to convert a binary ccp4 map file to a readable format so that I can retrieve the electron density at each real space grid point. Just tried MAPTONA4 and MAPEXCHANGE, but the resulting ascii file are not readable, and I didn't find any documentataion about how to read them. Could somebody give me any hint? Thanks a lot! Best Regards, Hailiang
Re: [ccp4bb] Structure Based Sequence Alignment
http://ww2.cs.mu.oz.au/~arun/Site/mustang.html http://tcoffee.vital-it.ch/cgi-bin/Tcoffee/tcoffee_cgi/index.cgi?stage1=1daction=EXPRESSO(3DCoffee)::Regular or maybe what you are searching for is Consurf ? http://consurf.tau.ac.il/ Jürgen On May 25, 2010, at 2:39 PM, Muhammed bashir Khan wrote: Dear All; Can some body tell me a website for structure based sequence alignment, which can also pin point the similar and identical residues in different colors. regards Bashir -- Muhammad Bashir Khan Department for Biomolecular Structural Chemistry Max F. Perutz Laboratories University of Vienna Campus Vienna Biocenter 5 A-1030 Vienna Austria Phone: +43(1)427752224 Fax: +43(1)42779522 - 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/
Re: [ccp4bb] Structure Based Sequence Alignment
I would also recommend STRAP if you haven't tried it ( http://www.bioinformatics.org/strap/) On Tue, May 25, 2010 at 2:53 PM, Jürgen Bosch jubo...@jhsph.edu wrote: http://ww2.cs.mu.oz.au/~arun/Site/mustang.html http://tcoffee.vital-it.ch/cgi-bin/Tcoffee/tcoffee_cgi/index.cgi?stage1=1daction=EXPRESSO(3DCoffee)::Regular or maybe what you are searching for is Consurf ? http://consurf.tau.ac.il/ Jürgen On May 25, 2010, at 2:39 PM, Muhammed bashir Khan wrote: Dear All; Can some body tell me a website for structure based sequence alignment, which can also pin point the similar and identical residues in different colors. regards Bashir -- Muhammad Bashir Khan Department for Biomolecular Structural Chemistry Max F. Perutz Laboratories University of Vienna Campus Vienna Biocenter 5 A-1030 Vienna Austria Phone: +43(1)427752224 Fax: +43(1)42779522 - 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/ http://web.me.com/bosch_lab/ -- Jim Fairman, Ph D. Post-Doctoral Fellow National Institutes of Health - NIDDK Cell: 1-865-748-8672 Lab: 1-301-594-9229 E-mail: fairman@gmail.com james.fair...@nih.gov
Re: [ccp4bb] how to read the ascii map converted by MAPTONA4 and MAPEXCHANGE
Hi, those formats are not intended to be readable other than by the format exchange programs, and I would strongly advise against trying! Instead, pretty well all programs read a CCP4 binary map directly and do whatever they need to with the map array in memory. The documentation for the Fortran API is in your $CDOC/maplib.doc and for the C++ API at (among others) http://www.ysbl.york.ac.uk/~cowtan/clipper/doc/p_develop_map.html . If you're really desperate you could probably hack it with the output from mapdump, though again that format was designed only for printing (in the days of the dim distant past when we contoured map sections by hand!), not for machine processing. Cheers -- Ian On Tue, May 25, 2010 at 7:49 PM, Hailiang Zhang zhan...@umbc.edu wrote: Hi, I wanted to convert a binary ccp4 map file to a readable format so that I can retrieve the electron density at each real space grid point. Just tried MAPTONA4 and MAPEXCHANGE, but the resulting ascii file are not readable, and I didn't find any documentataion about how to read them. Could somebody give me any hint? Thanks a lot! Best Regards, Hailiang
[ccp4bb] Catalytic residues in active sites
Hi there I wonder if anyone can recommend a good review/paper describing crystal structures that show high energy residues in active sites. By that I mean residues that may be in a strained conformation and rotate between conformations, such that they may even switch into unfavoured ramachandran regions during their activity, but that the strained high energy state is relevant to the enzyme activity? If I get enough responses I will list them and post them back up. Thanks so much in advance Gina Notice: This e-mail message, together with any attachments, contains information of Merck Co., Inc. (One Merck Drive, Whitehouse Station, New Jersey, USA 08889), and/or its affiliates Direct contact information for affiliates is available at http://www.merck.com/contact/contacts.html) that may be confidential, proprietary copyrighted and/or legally privileged. It is intended solely for the use of the individual or entity named on this message. If you are not the intended recipient, and have received this message in error, please notify us immediately by reply e-mail and then delete it from your system.
Re: [ccp4bb] Structure Based Sequence Alignment
On 26/05/2010, at 6:53 AM, Jürgen Bosch jubo...@jhsph.edu wrote: http://ww2.cs.mu.oz.au/~arun/Site/mustang.html We have built a web server for this at http://pxgrid.med.monash.edu.au:8080/mustangserver/ Cheers Ashley http://tcoffee.vital-it.ch/cgi-bin/Tcoffee/tcoffee_cgi/index.cgi?stage1=1daction=EXPRESSO(3DCoffee)::Regular or maybe what you are searching for is Consurf ? http://consurf.tau.ac.il/ Jürgen On May 25, 2010, at 2:39 PM, Muhammed bashir Khan wrote: Dear All; Can some body tell me a website for structure based sequence alignment, which can also pin point the similar and identical residues in different colors. regards Bashir -- Muhammad Bashir Khan Department for Biomolecular Structural Chemistry Max F. Perutz Laboratories University of Vienna Campus Vienna Biocenter 5 A-1030 Vienna Austria Phone: +43(1)427752224 Fax: +43(1)42779522 - 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/
Re: [ccp4bb] Is it possible for the Tris buffer to strip the Zn ions from the Zinc Finger motif of a protein?
Dear Heng, Tris forms only a very weak complex with Zn2+. It is unlikely that the problem arises from Tris. It might be simply that the complex dissociates in the presence of high imidazol when you elute the complex from the nickel column. Imidazol is a pretty good Zn2+ coordinating molecule. DTT in higher concentrations (1 mM should be now problem for a zinc finger) can also strip of a Zn2+; it is a high affinity bidental chelator. HTH Guenter Date: Sat, 22 May 2010 11:17:41 +0800 From: rh_ibp2...@hotmail.com Subject: [ccp4bb] To: CCP4BB@JISCMAIL.AC.UK Dear all, Recently, I am working on a complex which includes two protein subunits. The interaction was based on the Zinc Finger motif of one protein. I co-purified the complex by nickel affinity column with one protein bearing a C terminal His tag and the other without any affinity tags. However, the complex was disassociated when applied to size exclusion chromatography. The buffer I use for SEC is 20mM Tris-HCl, 150mM NaCl, 1mM DTT, 5% Glycerol, pH 7.5, whearas the buffer I use for nickel affinity column is 50mM Na2HPO4, 10mM KH2PO4, 137mM NaCl, 2.7mM KCl, 10% Glycerol, pH7.4. So I am wondering is it possible for the Tris buffer to strip the Zn ions from the Zinc Finger motif of one protein that leads to the destruction of the complex? I will be very appreciated if anyone has some experience in such case and would like to share with me! Sincerely, Heng Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China 搜索本应是彩色的,快来体验新一代搜索引擎-必应,精美图片每天换哦! 立即试 用！ http://cn.bing.com/?form=CRMADS 聊天+搜索+邮箱 想要轻松出游,手机MSN帮你搞定! 立刻下载！ http://3g.msn.cn/ -- *** Priv.Doz.Dr. Guenter Fritz Fachbereich Biologie Sektion Naturwissenschaften Universitaet Konstanz http://www.biologie.uni-konstanz.de/fritz Universitaetsstrasse 10 Postfach M665 D-78457 Konstanz e-mail: guenter.fr...@uni-konstanz.de Phone Office: +49-(0)7531 88 3205 Phone Lab : +49-(0)7531 88 3733 Fax: +49-(0)7531 88 2966
Re: [ccp4bb] Structure Based Sequence Alignment
Sorry, the link was broken (problem with tomcat not restarting properly): Use http://pxgrid.med.monash.edu.au/mustangmr-server cheers Ashley On 26/05/2010, at 8:08 AM, Dhirendra K Simanshu wrote: Hi It seems the link (the one you send) is not working! Can you resend the correct link! Thanks Dhirendra On Tue, May 25, 2010 at 4:05 PM, Ashley Buckle ashley.buc...@med.monash.edu.au wrote: On 26/05/2010, at 6:53 AM, Jürgen Bosch jubo...@jhsph.edu wrote: http://ww2.cs.mu.oz.au/~arun/Site/mustang.html We have built a web server for this at http://pxgrid.med.monash.edu.au:8080/mustangserver/ Cheers Ashley http://tcoffee.vital-it.ch/cgi-bin/Tcoffee/tcoffee_cgi/index.cgi?stage1=1daction=EXPRESSO(3DCoffee)::Regular or maybe what you are searching for is Consurf ? http://consurf.tau.ac.il/ Jürgen On May 25, 2010, at 2:39 PM, Muhammed bashir Khan wrote: Dear All; Can some body tell me a website for structure based sequence alignment, which can also pin point the similar and identical residues in different colors. regards Bashir -- Muhammad Bashir Khan Department for Biomolecular Structural Chemistry Max F. Perutz Laboratories University of Vienna Campus Vienna Biocenter 5 A-1030 Vienna Austria Phone: +43(1)427752224 Fax: +43(1)42779522 - 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/ -- Dhirendra K Simanshu Research Scholar Memorial Sloan-Kettering Cancer Center New York, NY, USA Associate Professor Ashley M Buckle NHMRC Senior Research Fellow The Department of Biochemistry and Molecular Biology, Faculty of Medicine Monash University, Clayton, Vic 3800 Australia http://www.med.monash.edu.au/biochem/staff/abuckle.html iChat/AIM: blindcaptaincat skype: ashley.buckle Tel: (613) 9902 9313 (office) Fax : (613) 9905 4699 mobile: 0430 913031
[ccp4bb] How large should the real space correlation coefficient be?
Hi, Have seen the real-space correlation used widely judging the map quality. Generally or empirically, in order to say an map (area) has good quality, how large should the real space correlation coefficient be? Say, is 0.8 good enough on a residue base? Any references about this will be greatly appreciated! Thanks! Best Regards, Hailiang
Re: [ccp4bb] How large should the real space correlation coefficient be?
On Tuesday 25 May 2010, Hailiang Zhang wrote: Hi, Have seen the real-space correlation used widely judging the map quality. Generally or empirically, in order to say an map (area) has good quality, how large should the real space correlation coefficient be? I do not think that the real space correlation coefficient is a measure of map quality per se. You could have an excellent experimental map but a lousy model and hence a poor correlation coefficient. Ethan Say, is 0.8 good enough on a residue base? Any references about this will be greatly appreciated! Thanks! Best Regards, Hailiang
[ccp4bb] How to calculate real-space CC by section?
Hi,
I am working on a real space correlation on a specif protein section using
CCP4 OVERLAPMAP. I am using the following scripts, not sure whether it is
good or not (didn't find in OVERLAPMAP documentation).
overlapmap \
mapin1 ${PDB}-1.map\
mapin2 ${PDB}-2.map\
mapin3 ${PDB}-mask.map \
eof
CORR SECT
CHAIN A $START $END
END
There is no error message, but the results make no difference no matter
how I change $START and $END. I am not sure whether the above script is
ok.
By the way, more importantly to me, if corr sect works at all, will it
print out a single CC value by integrating over the WHOLE region define by
the section range?
Thanks!
Best Regards, Hailiang
