Re: [ccp4bb] Proper Reference for Phil Evans Program POINTLESS
Hi Rebecca, The citation I use is from the 2005 CCP4 study weekend: Scaling and Assessment of Data Quality, Philip Evans, Acta Cryst D 62 72-82. It is open access from www.iucr.org. Cheers, Graeme From: CCP4 bulletin board [mailto:[EMAIL PROTECTED] On Behalf Of Page, Rebecca Sent: 10 July 2007 00:04 To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] Proper Reference for Phil Evans Program POINTLESS CCP4ers: What is the correct reference for Phil Evans Program POINTLESS? Thanks for your help! Rebecca
Re: [ccp4bb] restrained refinement in Refmac
It looks like as if your PDB is corrupt. For example there atoms B that should not be present inamino acids. Garib On 9 Jul 2007, at 19:55, JINJIN ZHANG wrote: Hello all, I'm working on a protein-DNA complex. My protein is a trimer and the crystal has 3 trimers in an AU. I used a pdb file including three subunits as a search model when I did molecular replacement in CCP4. I did a rigid body refinement in Recfmac the R-factor and R-free did not go down. Then I tried to do restrained refinement by using MTZ and PDB file ourput from MR but failed. The log file is attached at the bottom of this message. It looked like something wrong with the PDB file input. But I used the same PDB file to run rigid body refinement and did not get any error message. How can this happen? I sincerely appreciate any comment and suggestion. Best, Jinjin --- title of input coord file --- PDB_code: PDB_name: PDB_date:XX-XXX-XX WARNING : atom :OT1 ARG AA 226 is absent in the library ATTENTION: atom:OARG AA 226 is missing in the structure WARNING : atom :BSER BB -1 is absent in the library ATTENTION: atom:CSER BB -1 is missing in the structure WARNING : atom :BHIS BB 0 is absent in the library ATTENTION: atom:CHIS BB 0 is missing in the structure WARNING : atom :BMET BB 1 is absent in the library ATTENTION: atom:CMET BB 1 is missing in the structure WARNING : atom :BTHR BB 2 is absent in the library ATTENTION: atom:CTHR BB 2 is missing in the structure WARNING : atom :BPRO BB 3 is absent in the library ATTENTION: atom:CPRO BB 3 is missing in the structure WARNING : atom :BASP BB 4 is absent in the library ATTENTION: atom:CASP BB 4 is missing in the structure WARNING : atom :BILE BB 5 is absent in the library ATTENTION: atom:CILE BB 5 is missing in the structure WARNING : atom :BILE BB 6 is absent in the library ATTENTION: atom:CILE BB 6 is missing in the structure WARNING : atom :BLEU BB 7 is absent in the library ATTENTION: atom:CLEU BB 7 is missing in the structure WARNING : atom :BGLN BB 8 is absent in the library ATTENTION: atom:CGLN BB 8 is missing in the structure WARNING : atom :BARG BB 9 is absent in the library ATTENTION: atom:CARG BB 9 is missing in the structure WARNING : atom :BTHR BB 10 is absent in the library ATTENTION: atom:CTHR BB 10 is missing in the structure WARNING : atom :BGLY BB 11 is absent in the library ATTENTION: atom:CGLY BB 11 is missing in the structure WARNING : atom :BILE BB 12 is absent in the library ATTENTION: atom:CILE BB 12 is missing in the structure WARNING : atom :BASP BB 13 is absent in the library ATTENTION: atom:CASP BB 13 is missing in the structure WARNING : atom :BVAL BB 14 is absent in the library ATTENTION: atom:CVAL BB 14 is missing in the structure WARNING : atom :BARG BB 15 is absent in the library ATTENTION: atom:CARG BB 15 is missing in the structure WARNING : atom :BALA BB 16 is absent in the library ATTENTION: atom:CALA BB 16 is missing in the structure WARNING : atom :BVAL BB 17 is absent in the library ... and more ... ATTENTION: atom:CVAL BB 17 is missing in the structure ... and more ... Number of chains : 9 Total number of monomers :2052 Number of atoms : 17226 Number of missing atoms : 693 Number of rebuilt atoms : 0 Number of unknown atoms : 693 Number of deleted atoms : 0 Number of bonds restraints: 14889 Number of angles restraints : 18801 Number of torsions restraints :7878 Number of chiralities :1746 Number of planar groups :2244 IERR =1 There is error. See above ===> Error: Fatal error. Cannot continue Refmac_5.2.0019: Fatal error. Cannot continue Times: User: 0.0s System:0.0s Elapsed: 0:39 ** * * Information from CCP4Interface script ** * The program run with command: refmac5 XYZIN "C:/Ccp4Temp/ lam-6-14-07-P1/trimer_molrep1.pdb" XYZOUT "C:/Ccp4Temp/lam-6-14-07- P1/lam6-14_11_4_pdb_1.tmp" HKLIN "C:/Ccp4Temp/lam-6-14-07-P1/ ScalAveraged.mtz" HKLOUT "C:/Ccp4Temp/lam-6-14-07-P1/ lam6-14_11_6_mtz_1.tmp" LIBOUT "C:/Ccp4Temp/lam-6-14-07-P1/ lam6-14_11_lib.cif" has failed with error message Refmac_5.2.001
Re: [ccp4bb] Proper Reference for Phil Evans Program POINTLESS
Page, Rebecca escribió: CCP4ers: What is the correct reference for Phil Evans Program POINTLESS? Thanks for your help! Rebecca Evans, P. (2006) Scaling and assessment of data quality. /Acta Crystallogr. sect. D/, *62*, 72-82. --
Re: [ccp4bb] Proper Reference for Phil Evans Program POINTLESS
That's the right (& indeed only) one (although more & up-coming versions are a little different to that described there) Phil On 10 Jul 2007, at 08:36, Winter, G (Graeme) wrote: Hi Rebecca, The citation I use is from the 2005 CCP4 study weekend: Scaling and Assessment of Data Quality, Philip Evans, Acta Cryst D 62 72-82. It is open access from www.iucr.org. Cheers, Graeme From: CCP4 bulletin board [mailto:[EMAIL PROTECTED] On Behalf Of Page, Rebecca Sent: 10 July 2007 00:04 To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] Proper Reference for Phil Evans Program POINTLESS CCP4ers: What is the correct reference for Phil Evans Program POINTLESS? Thanks for your help! Rebecca
Re: [ccp4bb] Help with reducing crystal mosaicity
Mary, freezing habitually increases mosaicity. In your case, the high water content adds to the problem. Try not to freeze the crystal but collect at sub-zero temperature (in short glass capillaries or use oil plugs instead). You have to optimize the "close to freezing" data-collection temperature. I collected complete synchrotron datasets (of GCPII in buffer with PEG1500 and PEG400) at 260-263 Kelvin which resulted in mosaicity values of as small as 0.07 degrees! At 277 K, the crystals only last for a few images and freezing did not work (for the buffer mentioned before). - J. - -Original Message- From: Mary Fitzgerald <[EMAIL PROTECTED]> Date: Mon, 9 Jul 2007 18:05:10 To:CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] Help with reducing crystal mosaicity Help please! I'm looking for some new ideas. I have crystals that come out of a sitting drop with a mixture of sodium cacodylate at pH 6.5, magnesium acetate and MPD for the well solution. The MPD concentration is sufficient to act as a cryoprotectant. Currently, I directly freeze these crystals in liquid nitrogen. When I collect data, I typically have high anisotropic mosaicity; it ranges from 0.8 to 1.2. This is further complicated with a weakly diffracting crystal (4-5 A) that has a long unit cell axis of ~500 and often twinning. It has been suggested to me that the cryoprotectent is a problem. I haven't checked the diffraction at room temperature, yet. Please no suggestions of finding a different crystal form as that's not a consideration at the moment. I have my reasons. I did find one crystal that has lower mosaicity (0.5 to 0.8) but had weaker diffraction then the typical crystal. Attempts at flash cryoannealing have not helped. So, what's a good way to change the cryoprotectant if the cryoprotectant is the precipitant? I've considered trying dehydration but wasn't certain if that would help with the mosaicity. Thanks for any ideas, Mary X. Fitzgerald Postdoctoral Associate -- Jeroen Raymundus Mesters, Ph.D. Institut fuer Biochemie, Universitaet zu Luebeck Zentrum fuer Medizinische Struktur und Zellbiologie Ratzeburger Allee 160, D-23538 Luebeck Tel: +49-451-5004070, Fax: +49-451-5004068 E-mail: [EMAIL PROTECTED] Http://www.biochem.uni-luebeck.de Http://www.iobcr.org Http://www.opticryst.org -- If you can look into the seeds of time and say which grain will grow and which will not - speak then to me (Macbeth) --
[ccp4bb] Post Doc on Hsp90-Complexes at ICR, London
Postdoctoral Training Fellow Structural Biology of Hsp90 – Client Protein Complexes Section of Structural Biology The Institute of Cancer Research Chester Beatty Laboratories Chelsea, London The Institute of Cancer Research is one of the world's leading cancer research organisations and is internationally renowned for the quality of its science. Our mission is the relief of human suffering by pursuing excellence in the fight against cancer. A Wellcome Trust-funded Postdoctoral position is available from 1st October 2007 in Professor Laurence Pearl’s group in the Section of Structural Biology, to study the structural basis for regulation and activation of ‘client’ proteins by the Hsp90 molecular chaperone system. The Postdoc will be responsible for expression, purification, crystallization and structural analysis of protein complexes involving the Hsp90 molecular chaperone, co-chaperones and client proteins (such as protein kinases) that depend on Hsp90 for their biological function. The Section of Structural Biology at ICR is exceptionally well equipped for all aspects of modern structural biology, with state-of-the-art laboratories for molecular biology, recombinant expression in bacterial and eukaryotic systems, biochemistry, X-ray crystallography and electron microscopy. Applicants must have a PhD in a biological science, and experience in recombinant expression, protein purification, crystallisation and X-ray crystallography. Information on our previous work in this field can be found by searching PubMed and via our Home Page http://www.icr.ac.uk/structbi/ The starting salary for the position will be in the range £28,050 to £35,400 p.a. inclusive (based on previous post-doctoral experience) and the post is offered initially on a fixed term contract of up to 4 years. Informal enquiries to [EMAIL PROTECTED] Please DO NOT send your application to Professor Pearl; CVs must be submitted in line with the instructions below. To apply, please send two copies of your CV and covering letter (incl. the names and addresses of two referees) together with a completed Equal Opportunities Monitoring Form to the HR Office, The Institute of Cancer Research, 123 Old Brompton Road, London SW7 3RP quoting job Ref. C48. Closing date: 6 August 2007 -- Laurence H. Pearl PhD FMedSci Professor of Protein Crystallography and Section Co-Chairman Section of Structural Biology, Institute of Cancer Research Chester Beatty Laboratories, 237 Fulham Road, London SW3 6JB, UK Phone +44-(0)20 7153 5422 : Secretary +44-(0)20 7153 5443 FAX +44-(0)20 7153 5457 : E-Mail [EMAIL PROTECTED] -- " Live Simply and do Serious Things .. " - Dorothy Crowfoot Hodgkin -- The Institute of Cancer Research: Royal Cancer Hospital, a charitable Company Limited by Guarantee, Registered in England under Company No. 534147 with its Registered Office at 123 Old Brompton Road, London SW7 3RP. This e-mail message is confidential and for use by the addressee only. If the message is received by anyone other than the addressee, please return the message to the sender by replying to it and then delete the message from your computer and network.
Re: [ccp4bb] Help with reducing crystal mosaicity
You might also want to loook into using parathone for freezing... Or collection at 260K indeed! Flip -Original Message- From: CCP4 bulletin board [mailto:[EMAIL PROTECTED] On Behalf Of mesters Sent: Tuesday, July 10, 2007 11:01 To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] Help with reducing crystal mosaicity Mary, freezing habitually increases mosaicity. In your case, the high water content adds to the problem. Try not to freeze the crystal but collect at sub-zero temperature (in short glass capillaries or use oil plugs instead). You have to optimize the "close to freezing" data-collection temperature. I collected complete synchrotron datasets (of GCPII in buffer with PEG1500 and PEG400) at 260-263 Kelvin which resulted in mosaicity values of as small as 0.07 degrees! At 277 K, the crystals only last for a few images and freezing did not work (for the buffer mentioned before). - J. - > -Original Message- > From: Mary Fitzgerald <[EMAIL PROTECTED]> > Date: Mon, 9 Jul 2007 18:05:10 > To:CCP4BB@JISCMAIL.AC.UK > Subject: [ccp4bb] Help with reducing crystal mosaicity > > Help please! > > I'm looking for some new ideas. I have crystals that come out of a > sitting drop with a mixture of sodium cacodylate at pH 6.5, magnesium > acetate and MPD for the well solution. The MPD concentration is > sufficient to act as a cryoprotectant. Currently, I directly freeze > these crystals in liquid nitrogen. When I collect data, I typically > have high anisotropic mosaicity; it ranges from 0.8 to 1.2. This is > further complicated with a weakly diffracting crystal (4-5 A) that has > a long unit cell axis of ~500 and often twinning. > > It has been suggested to me that the cryoprotectent is a problem. I > haven't checked the diffraction at room temperature, yet. Please no > suggestions of finding a different crystal form as that's not a > consideration at the moment. I have my reasons. I did find one > crystal that has lower mosaicity (0.5 to 0.8) but had weaker > diffraction then the typical crystal. Attempts at flash cryoannealing > have not helped. > > So, what's a good way to change the cryoprotectant if the > cryoprotectant is the precipitant? I've considered trying dehydration > but wasn't certain if that would help with the mosaicity. > > Thanks for any ideas, > > Mary X. Fitzgerald > Postdoctoral Associate > -- Jeroen Raymundus Mesters, Ph.D. Institut fuer Biochemie, Universitaet zu Luebeck Zentrum fuer Medizinische Struktur und Zellbiologie Ratzeburger Allee 160, D-23538 Luebeck Tel: +49-451-5004070, Fax: +49-451-5004068 E-mail: [EMAIL PROTECTED] Http://www.biochem.uni-luebeck.de Http://www.iobcr.org Http://www.opticryst.org -- If you can look into the seeds of time and say which grain will grow and which will not - speak then to me (Macbeth) --
Re: [ccp4bb] Help with reducing crystal mosaicity
Hi Mary, There are many excellent suggestions already but you may want to try tweaking your cryo a bit. This is described very nicely by Ed Mitchell and Elspeth Garman in a paper on mosaic spread as a function of cryoprotectants concentration - J. Appl. Cryst. (1994) 27, 1070-1074. You already have a cryoprotectant, the MPD, and could up the concentration a little by successive transfer or adding a little to the drop. I've also found that (2R,3R)-(-)-2,3-butanediol works as a 'magic' elixir in very small concentrations when added directly to the drop. I'd say try room temperature but at that resolution you are going to need all the X-rays you can get onto it. Cheers, Eddie. Edward Snell Ph.D. Assistant Prof. Department of Structural Biology, SUNY Buffalo, Hauptman-Woodward Medical Research Institute 700 Ellicott Street, Buffalo, NY 14203-1102 Phone: (716) 898 8631 Fax: (716) 898 8660 Email: [EMAIL PROTECTED] Telepathy: 42.2 GHz Heisenberg was probably here! Mary, freezing habitually increases mosaicity. In your case, the high water content adds to the problem. Try not to freeze the crystal but collect at sub-zero temperature (in short glass capillaries or use oil plugs instead). You have to optimize the "close to freezing" data-collection temperature. I collected complete synchrotron datasets (of GCPII in buffer with PEG1500 and PEG400) at 260-263 Kelvin which resulted in mosaicity values of as small as 0.07 degrees! At 277 K, the crystals only last for a few images and freezing did not work (for the buffer mentioned before). - J. - > -Original Message- > From: Mary Fitzgerald <[EMAIL PROTECTED]> > Date: Mon, 9 Jul 2007 18:05:10 > To:CCP4BB@JISCMAIL.AC.UK > Subject: [ccp4bb] Help with reducing crystal mosaicity > > Help please! > > I'm looking for some new ideas. I have crystals that come out of a > sitting drop with a mixture of sodium cacodylate at pH 6.5, magnesium > acetate and MPD for the well solution. The MPD concentration is > sufficient to act as a cryoprotectant. Currently, I directly freeze > these crystals in liquid nitrogen. When I collect data, I typically > have high anisotropic mosaicity; it ranges from 0.8 to 1.2. This is > further complicated with a weakly diffracting crystal (4-5 A) that has > a long unit cell axis of ~500 and often twinning. > > It has been suggested to me that the cryoprotectent is a problem. I > haven't checked the diffraction at room temperature, yet. Please no > suggestions of finding a different crystal form as that's not a > consideration at the moment. I have my reasons. I did find one > crystal that has lower mosaicity (0.5 to 0.8) but had weaker > diffraction then the typical crystal. Attempts at flash cryoannealing > have not helped. > > So, what's a good way to change the cryoprotectant if the > cryoprotectant is the precipitant? I've considered trying dehydration > but wasn't certain if that would help with the mosaicity. > > Thanks for any ideas, > > Mary X. Fitzgerald > Postdoctoral Associate > -- Jeroen Raymundus Mesters, Ph.D. Institut fuer Biochemie, Universitaet zu Luebeck Zentrum fuer Medizinische Struktur und Zellbiologie Ratzeburger Allee 160, D-23538 Luebeck Tel: +49-451-5004070, Fax: +49-451-5004068 E-mail: [EMAIL PROTECTED] Http://www.biochem.uni-luebeck.de Http://www.iobcr.org Http://www.opticryst.org -- If you can look into the seeds of time and say which grain will grow and which will not - speak then to me (Macbeth) --
[ccp4bb] Beam Time at BCSB beamlines in Berkeley
== Synchrotron Beam Time available for Macromolecular Crystallography at Berkeley Center for Structural Biology Beamlines (5.0.1, 5.0.2, 5.0.3, 8.2.1 and 8.2.2). The ALS is now accepting General User applications for September/ October == The Berkeley Center for Structural Biology (BCSB) operates five beamlines at the Advanced Light Source in Berkeley. Beamlines 5.0.1, 5.0.2 and 5.0.3 are equipped with the Berkeley Automounter system. Beamlines 5.0.2, 5.0.3, 8.2.1 and 8.2.2 have large 3x3 Q315 or Q315r CCD detectors. The beamlines generate between 1.5x10^11 and 8x10^11 photons per second of flux into a 100 micron collimator. Please visit http://bcsb.lbl.gov/ for more details about the Center and its beamlines. If you'd like to apply for beamtime at these, or any other ALS macromolecular crystallography beamlines, please submit a General User proposal by July 15, 2007 in order to be considered for the September-October schedules. To find out more, click on: http://www-als.lbl.gov/als/quickguide/independinvest.html We invite you to submit a proposal at: http://alsusweb.lbl.gov/4DCGI/WEB_GetForm/PXProposalEntry.shtml/ Initialize. You can also request access to any open beam time on the five BCSB beamlines by e-mailing our scheduler at [EMAIL PROTECTED] Any industrial or academic group interested in becoming part of the Participating Research Team for beamlines 5.0.1 and 5.0.2 should contact the Head of the BCSB (Paul Adams: [EMAIL PROTECTED], 510-486-4225). (Please note that executed user agreements and proprietary fees, if applicable, must be received at least five working days prior to scheduled beam time.) -- Paul Adams Senior Staff Scientist, Physical Biosciences Division Head, Berkeley Center for Structural Biology Deputy Principal Investigator, Berkeley Structural Genomics Center Building 64, Room 248 Tel: 510-486-4225, Fax: 510-486-5909 http://cci.lbl.gov/ Lawrence Berkeley Laboratory 1 Cyclotron Road BLDG 64R0121 Berkeley, CA 94720, USA. --
Re: [ccp4bb] Help with reducing crystal mosaicity
Jeroen brings up a good point. Back in the old days, around 5 B. C. (Before Cryo), we would use a chilled air generator to blow a stream of cold air along the capillary axis to keep the crystals just above their freezing point--it made a huge difference in crystal lifetime. I recall a colleague devising an apparatus from a 50 ml conical tube. The bottom was cut off and cold air was blown in from the other end. Windows were cut in either side to allow the beam to pass & covered in mylar. This way the entire capillary was contained within the cold tube, so no temperature gradients formed along the length of the capillary (temp gradient => distillation => dead crystal). Later, we purchased a very clever goniometer head from Nonius that had a plastic cylinder attached to goniometer head, with a swivel, so the hose supplying cold air didn't get tangled during data collection... I've often thought duplicating this apparatus when we encounter cryo problems, but I'm always stymied when trying to find a cheap and simple source of cold air. Any bright ideas? On Jul 10, 2007, at 5:00 AM, mesters wrote: Mary, freezing habitually increases mosaicity. In your case, the high water content adds to the problem. Try not to freeze the crystal but collect at sub-zero temperature (in short glass capillaries or use oil plugs instead). You have to optimize the "close to freezing" data-collection temperature. I collected complete synchrotron datasets (of GCPII in buffer with PEG1500 and PEG400) at 260-263 Kelvin which resulted in mosaicity values of as small as 0.07 degrees! At 277 K, the crystals only last for a few images and freezing did not work (for the buffer mentioned before). - J. - --- Patrick J. Loll, Ph. D. (215) 762-7706 Associate Professor FAX: (215) 762-4452 Department 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 [EMAIL PROTECTED]
Re: [ccp4bb] Help with reducing crystal mosaicity
CCP4 bulletin board wrote on 07/10/2007 02:07:37 PM: > Jeroen brings up a good point. Back in the old days, around 5 B. C. > (Before Cryo), we would use a chilled air generator to blow a stream > of cold air along the capillary axis to keep the crystals just above > their freezing point--it made a huge difference in crystal lifetime. > I recall a colleague devising an apparatus from a 50 ml conical > tube. The bottom was cut off and cold air was blown in from the > other end. Windows were cut in either side to allow the beam to pass > & covered in mylar. This way the entire capillary was contained > within the cold tube, so no temperature gradients formed along the > length of the capillary (temp gradient => distillation => dead > crystal). Later, we purchased a very clever goniometer head from > Nonius that had a plastic cylinder attached to goniometer head, with > a swivel, so the hose supplying cold air didn't get tangled during > data collection... > > I've often thought duplicating this apparatus when we encounter cryo > problems, but I'm always stymied when trying to find a cheap and > simple source of cold air. Any bright ideas? > Hi Patrick - Many cryosystems (definitely the Cryojet, and I believe the Cryostream) can be set to run at any temperature between room temp and liquid nitrogen temp. I'm not sure of the temperature stability at temps > 0 C, and you might burn out the heaters prematurely if you do this all the time, but it should work. Then you just need to move the nozzle so it's coaxial with your goniostat's rotation axis, and aim the capillary down the nozzle. You could probably even move the capillary *inside* the cryo nozzle a bit, so only the bit with the crystal is in the free stream. It's not a cheap solution, but you've almost certainly got a cryosystem already... - Matt -- Matthew Franklin , Ph.D. Senior Scientist, ImClone Systems 180 Varick Street, 6th floor New York, NY 10014 phone:(917)606-4116 fax:(212)645-2054 Confidentiality Note: This e-mail, and any attachment to it, contains privileged and confidential information intended only for the use of the individual(s) or entity named on the e-mail. If the reader of this e-mail is not the intended recipient, or the employee or agent responsible for delivering it to the intended recipient, you are hereby notified that reading it is strictly prohibited. If you have received this e-mail in error, please immediately return it to the sender and delete it from your system. Thank you.
[ccp4bb] shelx HKLF4 to mtz conversion?
I have a Shelx HKLF4 formated reflection file that I would like to convert to a mtz file. The primary reason is to transfer the freeR set. I found a couple of convoluted ways of converting the reflection data, but no real way of getting the freeR set to transfer. Any suggestions? Cheers Leonard M. Thomas Ph.D. Director, Macromolecular Crystallography Laboratory Howard Hughes Medical Institute California Institute of Technology Division of Biology 1200 E. California Blvd. MC 114-96 Pasadena, CA 91125 626-395-2453 [EMAIL PROTECTED] http://www.br.caltech.edu/cmclab
[ccp4bb] Postdoctoral Fellow position
This notification is posted on behalf of Prof Wim Hol. Interested parties should reply to him at: [EMAIL PROTECTED] or at the address below. Postdoctoral Fellow Structural Biology of the RNA editing editosome JOB DESCRIPTION: The project aims at understanding the structure and functioning of the unique RNA-editing editosome from Trypanosoma brucei and related trypanosomatid protozoan species. These organisms are the causative agents of a variety of diseases in tropical and subtropical areas, including sleeping sickness in Sub-Saharan Africa, Chagas disease in Latin America and leishmaniasis throughout the tropics and subtropics. Several essential genes in the mitochondria of these protozoa undergo a fascinating U-insertion/deletion RNA editing process involving several protein and multi-protein complexes. The RNA-editing editosome of about 1.5 million Daltons performs the actual editing steps, involving an enzyme-cascade principle. The editosome consists of over 15 different proteins with most, if not all, of these proteins present in multiple copies. The goal of the project is to unravel, by molecular biology and crystallographic approaches: (i) protein-protein and protein-RNA interactions within the editosome; and, (ii) crystal structures of components and sub-complexes of this multi-protein assembly in complex with a variety of RNA molecules. Since several of the editosome proteins have been shown to be essential for the parasites, the results obtained provide a platform for structure-based drug design. The successful candidate will have the opportunity to carry out (i) molecular biology, protein expression and purification methods to obtain insight into protein-protein and protein-RNA interactions involving the editosome, and (ii) to determine high resolution crystal structures of individual editosome components, and in particular of large sub-complexes of the editosome with RNA. Numerous expression systems are already available for preparing complexes containing multiple editosome proteins. For further information regarding the editosome see: Deng, J., Schnaufer, A., Salavati, R., Stuart, K. & Hol, W. G. J. (2004). High Resolution Crystal Structure of an Editosome Enzyme from Trypanosoma brucei: RNA Editing ligase I. J Mol Biol 343, 601-613 Deng, J., Lewis Ernst, N., Turley, S., Stuart, K. & Hol, W. G. J. (2005). Structural basis for UTP specificity of RNA editing TUTases from Trypanosoma brucei. EMBO J. 24, 4007-4017. For information regarding the research in our lab see the websites: http://www.bmsc.washington.edu/WimHol/ and http://depts.washington.edu/biowww/faculty/hol.html JOB REQUIREMENTS: - Good knowledge and extensive experience with molecular biology for protein overexpression - Experience with: - protein expression and purification methods of soluble proteins in E coli - characterizing purified soluble proteins - crystallization methods for proteins - protein crystal cryo-protection procedures - protein structure determination methods, including: X-ray diffraction data collection and data processing; selenomethionine SAD and MAD phasing and/or multiple isomorphous replacement and/or molecular replacement; density modification; model building; crystallographic refinement; structure analysis and structure validation procedures using multiple computer programs and interactive graphics techniques. - Excellent interpersonal skills to function optimally in the editosome project team and to cooperate with collaborators in other institutions. The Following experience would be a plus: - Molecular biology for obtaining truncated protein variants and implementing surface mutations to enhance the probability of crystal growth - production of RNA - protein expression and purification methods of soluble proteins in insect cells START DATE: Immediately INSTITUTION:Department of Biochemistry Biomolecular Structure Center School of Medicine Box 357742 University of Washington Seattle, WA, 98195 USA
Re: [ccp4bb] shelx HKLF4 to mtz conversion?
You can use f2mtz with the format identifier to convert the file: 8< snip-> script start <-- #/bin/bash f2mtz hklin yourshelxfile.hkl hklout yourmtzfile.mtz << eof cell"you cell dimensions" symm "your space group" format '(3F4.0,2F8.2,F4.0)' ctypout H H H J Q R LABOUT H K L Intensity stddevIntensity freeR end eof --->script end <---snap >8 The important bits are the format line and the ctypout line, and you have to add your cell and symmetry. That can probably even be done through the gui, but a script is simpler (to me, anyway). Hope this helps, Tim On Wednesday 11 July 2007 09:29, Leonard Thomas wrote: > I have a Shelx HKLF4 formated reflection file that I would like to > convert to a mtz file. The primary reason is to transfer the freeR > set. I found a couple of convoluted ways of converting the > reflection data, but no real way of getting the freeR set to transfer. > > Any suggestions? > > Cheers > > > Leonard M. Thomas Ph.D. > Director, Macromolecular Crystallography Laboratory > Howard Hughes Medical Institute > California Institute of Technology > Division of Biology > 1200 E. California Blvd. MC 114-96 > Pasadena, CA 91125 > 626-395-2453 > [EMAIL PROTECTED] > http://www.br.caltech.edu/cmclab -- Tim Grune Australian Synchrotron 800 Blackburn Road Clayton, VIC 3168 Australia pgpuMHpGIBBuY.pgp Description: PGP signature
Re: [ccp4bb] Pymol, labels, maps
Hi, I like to short out how I can see map in pymol and bobscript. When I am using pymol, I cann't see map file. Same map file I can see in Coot. Map file (created for O using from .fcf shelxpro) is also not coming; extenstion .xplor (or .dsn6) not working either. message: Crystal: Unit Cell Volume 845180. ExecutiveLoad: "popp_fcf_sigmaa1_FP1_map.ccp4" loaded as "popp_fcf_sigmaa1_FP1_map", through state 0. How one can label symbols in pymols. Example shown at "http://www.pymolwiki.org/index.php/Label"; does not give symbolic label. In bobscript when I use map file (created using shelxpro for O format) as per the example shown in "http://www.strubi.ox.ac.uk/bobscript/example3.html"; error message comes. message: . contouring map shelx at 4.00 Error: no graphical segments to output end_plot line number 36 Maximum actual usage of allocated memory: 60606 out of 120 "map" points used (leaving 4.3 Mb unused) Line number 36 is "end_plot". I am not sure why is this error for, Appreciate suggestion and help. Thanks Sam _ PC Magazine’s 2007 editors’ choice for best web mail—award-winning Windows Live Hotmail. http://imagine-windowslive.com/hotmail/?locale=en-us&ocid=TXT_TAGHM_migration_HMWL_mini_pcmag_0707
