Re: [ccp4bb] Symmetry problem
-BEGIN PGP SIGNED MESSAGE- Hash: SHA1 Dear Monika, What program did you use for MR? I would expect the output PDB file has the correct cell and symmetry from the mtz-file you replaced against. To answer your question: you can simply concatenate the two files (e.g. with a text editor), remove the CRYST1 card and set it with pdbset: pdbset xyzin your.pdb xyzout yourpdb_with_new_sg.pdb eof CELL yourA yourB yourC yourAlpha yourBeta yourGamma SPAC yourSpacegroup end eof Best, Tim On 02/18/2014 06:59 PM, Monika Coronado wrote: Dear, Does anyone know how to merge two molecules with different symmetry? I will explain: I have done the molecular replacement using the domains of the molecules separately, now I have to put all together, however they have a different symmetry. I will appreciate any kind of help. Regards, Mnika - -- - -- Dr Tim Gruene Institut fuer anorganische Chemie Tammannstr. 4 D-37077 Goettingen GPG Key ID = A46BEE1A -BEGIN PGP SIGNATURE- Version: GnuPG v1.4.12 (GNU/Linux) Comment: Using GnuPG with Icedove - http://www.enigmail.net/ iD8DBQFTBHhtUxlJ7aRr7hoRApb1AJ9FwxpTIbtcIlJyuU0Rn7u1UxLHrACglchR weQEKA+/lnmIkQKtHd+nhj4= =LgoX -END PGP SIGNATURE-
Re: [ccp4bb] Off topic Chromatography systems
Agilent also has some decent systems. Cheers, Ronnie -- Ronnie Berntsson, PhD PostDoctoral Fellow Department of Biochemistry and Biophysics Arrhenius Laboratories for Natural Sciences Stockholm University 10691 Stockholm Sweden On 18 Feb 2014, at 17:15 , Scott Pegan scott.d.pe...@gmail.com wrote: Hey everyone, I am looking at acquiring two new FPLC chromatography systems for purification of protein to meet crystallographic needs. Naturally I am looking into Akta and have noticed Biorad's NGC systems. Are there any others out there right now? Scott -- Scott D. Pegan, Ph.D. Assistant Professor Chemistry Biochemistry University of Denver Office: 303 871 2533 Fax: 303 871 2254
Re: [ccp4bb] Ask for recommendation: crystal screening kit
Dear Wenhe, shameless plug You could also consider one of the screens available from Rigaku Reagents, former Emerald Bio (http://www.rigakureagents.com/). These include the Classic Wizard Screens, the JCSG+ and PEG Ion screens as well as screens for LCP and a Membrane Protein Extraction Detergent Screening Kit. / shameless plug Good luck! Best wishes, Bram ___ Bram Schierbeek | Senior App. Sci. | Rigaku Europe | bram.schierb...@rigaku.com| M: +31 6 13 989 469 Date:Mon, 17 Feb 2014 23:37:37 +0800 From:Wenhe wenhezhong.xmu@gmail.com Subject: Ask for recommendation: crystal screening kit Dear CCP4 friends, Sorry first for the out of topic request. We are going to buy more crystal screening kit, however, we are only familiar with some popular kits which we already have in the lab (listing below). Do you have your favourite kits which can share with us? Our lab more focus on enzymes and DNA/RNA binding proteins. Thank you. The kits we have in the lab: 1. Crystal Screen 1 and 2 (Hampton Research) 2. Index (Hampton Research) 3. Natrix 1 and 2 (Hampton Research) 4. PegRx 1 and 2 (Hampton Research) 5. Peg-Ion 1 and 2 (Hampton Research) 6. JCSG+ (Molecular Dimensions) 7. Morpheus (Molecular Dimensions) 8. Midas (Molecular Dimensions) Kind regards, Wenhe
[ccp4bb] AW: [ccp4bb] Symmetry problem
Another option would be the merge molecules option in coot (calculate - merge molecules). In coot you would also be able to move the molecules to the same asymmetric unit if that would be necessary. However, depending on the space group the MR solutions could have different origins and with polar space groups one (or three in P1!) coordinates may be arbitrary, prohibiting any merging efforts. The best way to go is to search for both molecules in one go. E.g. Phaser has the possibility to give two or more search models and then first search with the first model, and then with the second in the context of the solution for the first solution. I am sure other MR programs have similar options. If you wish, I could provide you with a sample command file. Best regards, Herman -Ursprüngliche Nachricht- Von: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] Im Auftrag von Tim Gruene Gesendet: Mittwoch, 19. Februar 2014 10:25 An: CCP4BB@JISCMAIL.AC.UK Betreff: Re: [ccp4bb] Symmetry problem -BEGIN PGP SIGNED MESSAGE- Hash: SHA1 Dear Monika, What program did you use for MR? I would expect the output PDB file has the correct cell and symmetry from the mtz-file you replaced against. To answer your question: you can simply concatenate the two files (e.g. with a text editor), remove the CRYST1 card and set it with pdbset: pdbset xyzin your.pdb xyzout yourpdb_with_new_sg.pdb eof CELL yourA yourB yourC yourAlpha yourBeta yourGamma SPAC yourSpacegroup end eof Best, Tim On 02/18/2014 06:59 PM, Monika Coronado wrote: Dear, Does anyone know how to merge two molecules with different symmetry? I will explain: I have done the molecular replacement using the domains of the molecules separately, now I have to put all together, however they have a different symmetry. I will appreciate any kind of help. Regards, Mnika - -- - -- Dr Tim Gruene Institut fuer anorganische Chemie Tammannstr. 4 D-37077 Goettingen GPG Key ID = A46BEE1A -BEGIN PGP SIGNATURE- Version: GnuPG v1.4.12 (GNU/Linux) Comment: Using GnuPG with Icedove - http://www.enigmail.net/ iD8DBQFTBHhtUxlJ7aRr7hoRApb1AJ9FwxpTIbtcIlJyuU0Rn7u1UxLHrACglchR weQEKA+/lnmIkQKtHd+nhj4= =LgoX -END PGP SIGNATURE-
Re: [ccp4bb] High Salt Cryo
For crystals grown out of a 2 uL drop of 1.2-1.8 M LiSO4 or 1.6-2.4 M AmmSO4, we do in situ cryoprotection with sodium malonate. We add 2-4 uL of 1.9 M Na malonate to the crystallization drop, wait 10 seconds and add 2-4 uL of 2.4 M sodium malonate, repeat with 2.8 M and then 3.4 M. We do not bother withdrawing aliquots to maintain a fixed volume. You may need to tweak the volumes to optimize the resulting diffraction. You can also break the additions at given concentration into smaller aliquots to reduce the osmotic shock. This approach is much gentler than transferring the crystal directly to 3 M sodium malonate. Do not leave the drop exposed to the air for more than 3 minutes or so because salt crystals will start to grow. When there are multiple crystals in a drop, often the unused crystals in the very high salt solution will still diffract well up to a year later if the crystallization chamber is resealed well; their diffraction might even improve with the prolonged exposure to high salt. Blaine Mooers Assistant Professor Department of Biochemistry and Molecular Biology University of Oklahoma Health Sciences Center S.L. Young Biomedical Research Center Rm. 466 Shipping address: 975 NE 10th Street, BRC 466 Oklahoma City, OK 73104-5419 Letter address: P.O. Box 26901, BRC 466 Oklahoma City, OK 73190 office: (405) 271-8300 lab: (405) 271-8313 fax: (405) 271-3910 e-mail: blaine-moo...@ouhsc.edu Faculty webpage: http://www.oumedicine.com/department-of-biochemistry-and-molecular-biology/faculty/blaine-mooers-ph-d- X-ray lab webpage: http://www.oumedicine.com/department-of-biochemistry-and-molecular-biology/department-facilities/macromolecular-crystallography-laboratory Small Angle Scattering webpage: http://www.oumedicine.com/docs/default-source/ad-biochemistry-workfiles/small-angle-scattering-links.html?sfvrsn=0 From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] On Behalf Of Katherine Sippel [katherine.sip...@gmail.com] Sent: Tuesday, February 18, 2014 12:08 PM To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] High Salt Cryo Hi all, I'm looking for a cryo condition for high NaCl (3+ M) crystallization condition. I would do it the proper way, but our beam/cryostream is down. I've tried a bunch of things at the moment. Ethylene glycol and PEG 400 nuke the crystals immediately even at low concentrations. Prolonged exposure to glycerol and sucrose starts to break them down so I'm thinking that the diffraction will probably suffer. I can't find any reports of NaCl's viability as a cryosalt. I've got Paratone/Paraffin oil/Mitegen's LV cryo oil on tap but I was hoping to not put all my eggs in one basket. I tried the ISRDB database through archive.comhttps://urldefense.proofpoint.com/v1/url?u=http://archive.comk=7DHVT22D9IhC0F3WohFMBA%3D%3D%0Ar=ftLbjJYpc5s5JQz9Q6qd7uT7FxPLb4V0aIwH4RJhyZU%3D%0Am=Vjr4m%2Fds%2FdLGOVQoQ0x8PApF%2FzyGkSwsbIoq92CSnOk%3D%0As=3cfbf18821b5b59934971bf583cf3dd6e2ded91923c614e670857e10916c687e without any luck (no search function). I've gone to the PDB searching for similar crystallization conditions and looked up the papers for their cryos, but they are all glycerol. Google gives me the same. I thought I'd see if anyone on the bb has an anecdotal this worked for us story. I would love to hear it. Thank you for your time, Katherine -- Nil illegitimo carborundum - Didactylos
Re: [ccp4bb] I/sigmaI or I/sigmaI
Dear Richard Gillilan, Where to find the archives of Dec 2003? I can only find the archives until 2007 at jiscmail. Thanks. Regards, Qixu Cai 发件人: Richard Gillilan r...@cornell.edu 答复: Richard Gillilan r...@cornell.edu 日期: 2014年2月16日 星期日 上午12:19 至: CCP4BB@JISCMAIL.AC.UK 主题: Re: [ccp4bb] I/sigmaI or I/sigmaI There was an informative discussion on this very topic back in Dec 1-2, 2003 if you browse the CCP4BB archives. Richard Gillilan MacCHESS On Feb 12, 2014, at 6:43 AM, Cai Qixu wrote: Dear all, Does the I/sigmaI in “Table 1” mean for I/sigmaI or I/sigmaI ? Thanks for your answer. Best wishes, Qixu Cai
Re: [ccp4bb] I/sigmaI or I/sigmaI
Dear Qixu Cai, You can find information about where to find the archives here: http://www.ccp4.ac.uk/ccp4bb.php#archives Best regards, Folmer 2014-02-19 14:44 GMT+01:00 Cai Qixu caiq...@gmail.com: Dear Richard Gillilan, Where to find the archives of Dec 2003? I can only find the archives until 2007 at jiscmail. Thanks. Regards, Qixu Cai 发件人: Richard Gillilan r...@cornell.edu 答复: Richard Gillilan r...@cornell.edu 日期: 2014年2月16日 星期日 上午12:19 至: CCP4BB@JISCMAIL.AC.UK 主题: Re: [ccp4bb] I/sigmaI or I/sigmaI There was an informative discussion on this very topic back in Dec 1-2, 2003 if you browse the CCP4BB archives. Richard Gillilan MacCHESS On Feb 12, 2014, at 6:43 AM, Cai Qixu wrote: Dear all, Does the I/sigmaI in “Table 1” mean for I/sigmaI or I/sigmaI ? Thanks for your answer. Best wishes, Qixu Cai -- Folmer Fredslund
Re: [ccp4bb] High Salt Cryo
4M NaCl should work too. It worked for the conditions with 1.8 - 2.0 M NaCl. Karolina W dniu 2014-02-19 06:38, Mooers, Blaine H.M. (HSC) napisał(a): For crystals grown out of a 2 uL drop of 1.2-1.8 M LiSO4 or 1.6-2.4 M AmmSO4, we do in situ cryoprotection with sodium malonate. We add 2-4 uL of 1.9 M Na malonate to the crystallization drop, wait 10 seconds and add 2-4 uL of 2.4 M sodium malonate, repeat with 2.8 M and then 3.4 M. We do not bother withdrawing aliquots to maintain a fixed volume. You may need to tweak the volumes to optimize the resulting diffraction. You can also break the additions at given concentration into smaller aliquots to reduce the osmotic shock. This approach is much gentler than transferring the crystal directly to 3 M sodium malonate. Do not leave the drop exposed to the air for more than 3 minutes or so because salt crystals will start to grow. When there are multiple crystals in a drop, often the unused crystals in the very high salt solution will still diffract well up to a year later if the crystallization chamber is resealed well; their diffraction might even improve with the prolonged exposure to high salt. Blaine Mooers Assistant Professor Department of Biochemistry and Molecular Biology University of Oklahoma Health Sciences Center S.L. Young Biomedical Research Center Rm. 466 Shipping address: 975 NE 10th Street, BRC 466 Oklahoma City, OK 73104-5419 Letter address: P.O. Box 26901, BRC 466 Oklahoma City, OK 73190 office: (405) 271-8300 lab: (405) 271-8313 fax: (405) 271-3910 e-mail: blaine-moo...@ouhsc.edu Faculty webpage: http://www.oumedicine.com/department-of-biochemistry-and-molecular-biology/faculty/blaine-mooers-ph-d- [1] X-ray lab webpage: http://www.oumedicine.com/department-of-biochemistry-and-molecular-biology/department-facilities/macromolecular-crystallography-laboratory [2] Small Angle Scattering webpage: http://www.oumedicine.com/docs/default-source/ad-biochemistry-workfiles/small-angle-scattering-links.html?sfvrsn=0 [3] From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] On Behalf Of Katherine Sippel [katherine.sip...@gmail.com] Sent: Tuesday, February 18, 2014 12:08 PM To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] High Salt Cryo Hi all, I'm looking for a cryo condition for high NaCl (3+ M) crystallization condition. I would do it the proper way, but our beam/cryostream is down. I've tried a bunch of things at the moment. Ethylene glycol and PEG 400 nuke the crystals immediately even at low concentrations. Prolonged exposure to glycerol and sucrose starts to break them down so I'm thinking that the diffraction will probably suffer. I can't find any reports of NaCl's viability as a cryosalt. I've got Paratone/Paraffin oil/Mitegen's LV cryo oil on tap but I was hoping to not put all my eggs in one basket. I tried the ISRDB database through archive.comhttps://urldefense.proofpoint.com/v1/url?u=http://archive.comk=7DHVT22D9IhC0F3WohFMBA%3D%3D%0Ar=ftLbjJYpc5s5JQz9Q6qd7uT7FxPLb4V0aIwH4RJhyZU%3D%0Am=Vjr4m%2Fds%2FdLGOVQoQ0x8PApF%2FzyGkSwsbIoq92CSnOk%3D%0As=3cfbf18821b5b59934971bf583cf3dd6e2ded91923c614e670857e10916c687e [4] without any luck (no search function). I've gone to the PDB searching for similar crystallization conditions and looked up the papers for their cryos, but they are all glycerol. Google gives me the same. I thought I'd see if anyone on the bb has an anecdotal this worked for us story. I would love to hear it. Thank you for your time, Katherine -- Nil illegitimo carborundum - Didactylos Links: -- [1] http://www.oumedicine.com/department-of-biochemistry-and-molecular-biology/faculty/blaine-mooers-ph-d- [2] http://www.oumedicine.com/department-of-biochemistry-and-molecular-biology/department-facilities/macromolecular-crystallography-laboratory [3] http://www.oumedicine.com/docs/default-source/ad-biochemistry-workfiles/small-angle-scattering-links.html?sfvrsn=0 [4] https://urldefense.proofpoint.com/v1/url?u=http://archive.comamp;k=7DHVT22D9IhC0F3WohFMBA%3D%3D%0Aamp;r=ftLbjJYpc5s5JQz9Q6qd7uT7FxPLb4V0aIwH4RJhyZU%3D%0Aamp;m=Vjr4m%2Fds%2FdLGOVQoQ0x8PApF%2FzyGkSwsbIoq92CSnOk%3D%0Aamp;s=3cfbf18821b5b59934971bf583cf3dd6e2ded91923c614e670857e10916c687e
Re: [ccp4bb] [ccp4bb] Symmetry problem
Hi, Monika, It sounds like you got the two domains separately in the asymmetric unit. You could do the symmetric operation for one of the domains in pymol by generating the symmetric mates (say 10 Å). Then you should see several same ones there. Select the one you need, and save the coordinates, then you can combine this to the other domain. Cheers, Shu --- Shu Xu, Ph.D. Research Fellow Department of Biochemistry Vanderbilt University School of Medicine Tel: (615)-343-7327 E-mail: xushuh...@gmail.com On Feb 19, 2014, at 4:40 AM, herman.schreu...@sanofi.com wrote: Another option would be the merge molecules option in coot (calculate - merge molecules). In coot you would also be able to move the molecules to the same asymmetric unit if that would be necessary. However, depending on the space group the MR solutions could have different origins and with polar space groups one (or three in P1!) coordinates may be arbitrary, prohibiting any merging efforts. The best way to go is to search for both molecules in one go. E.g. Phaser has the possibility to give two or more search models and then first search with the first model, and then with the second in the context of the solution for the first solution. I am sure other MR programs have similar options. If you wish, I could provide you with a sample command file. Best regards, Herman -Ursprüngliche Nachricht- Von: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] Im Auftrag von Tim Gruene Gesendet: Mittwoch, 19. Februar 2014 10:25 An: CCP4BB@JISCMAIL.AC.UK Betreff: Re: [ccp4bb] Symmetry problem -BEGIN PGP SIGNED MESSAGE- Hash: SHA1 Dear Monika, What program did you use for MR? I would expect the output PDB file has the correct cell and symmetry from the mtz-file you replaced against. To answer your question: you can simply concatenate the two files (e.g. with a text editor), remove the CRYST1 card and set it with pdbset: pdbset xyzin your.pdb xyzout yourpdb_with_new_sg.pdb eof CELL yourA yourB yourC yourAlpha yourBeta yourGamma SPAC yourSpacegroup end eof Best, Tim On 02/18/2014 06:59 PM, Monika Coronado wrote: Dear, Does anyone know how to merge two molecules with different symmetry? I will explain: I have done the molecular replacement using the domains of the molecules separately, now I have to put all together, however they have a different symmetry. I will appreciate any kind of help. Regards, Mnika - -- - -- Dr Tim Gruene Institut fuer anorganische Chemie Tammannstr. 4 D-37077 Goettingen GPG Key ID = A46BEE1A -BEGIN PGP SIGNATURE- Version: GnuPG v1.4.12 (GNU/Linux) Comment: Using GnuPG with Icedove - http://www.enigmail.net/ iD8DBQFTBHhtUxlJ7aRr7hoRApb1AJ9FwxpTIbtcIlJyuU0Rn7u1UxLHrACglchR weQEKA+/lnmIkQKtHd+nhj4= =LgoX -END PGP SIGNATURE-
Re: [ccp4bb] High Salt Cryo
Dear All, I would like to point out that the conditions 1.8 - 2.0 M NaCl are not considered High Salt as NaCl is soluble to 5M and a 2X solution (i.e. 4M NaCl) is possible. Also NaCl contrary to ammonium sulfate, citrate, phosphate, etc. is compatible with polyethylene glycol without phase separation problems. This means that with 1.8 - 2.0 M NaCl you have an vast repertoire of possible ways to cryo-protect crystals and with the vast repertoire you gain a good possibility of finding conditions that enhance diffraction: see: Vera, L., Stura, E. A. (2013) Strategies for protein cryocrystallography. Crystal Growth Design, http://pubs.acs.org/doi/full/10.1021/cg301531f For me the definition for High Salt is that 2X for the precipitant component is not possible. Enrico. On Wed, 19 Feb 2014 16:06:27 +0100, Karolina Michalska dzi...@amu.edu.pl wrote: 4M NaCl should work too. It worked for the conditions with 1.8 - 2.0 M NaCl. Karolina W dniu 2014-02-19 06:38, Mooers, Blaine H.M. (HSC) napisał(a): For crystals grown out of a 2 uL drop of 1.2-1.8 M LiSO4 or 1.6-2.4 M AmmSO4, we do in situ cryoprotection with sodium malonate. We add 2-4 uL of 1.9 M Na malonate to the crystallization drop, wait 10 seconds and add 2-4 uL of 2.4 M sodium malonate, repeat with 2.8 M and then 3.4 M. We do not bother withdrawing aliquots to maintain a fixed volume. You may need to tweak the volumes to optimize the resulting diffraction. You can also break the additions at given concentration into smaller aliquots to reduce the osmotic shock. This approach is much gentler than transferring the crystal directly to 3 M sodium malonate. Do not leave the drop exposed to the air for more than 3 minutes or so because salt crystals will start to grow. When there are multiple crystals in a drop, often the unused crystals in the very high salt solution will still diffract well up to a year later if the crystallization chamber is resealed well; their diffraction might even improve with the prolonged exposure to high salt. Blaine Mooers Assistant Professor Department of Biochemistry and Molecular Biology University of Oklahoma Health Sciences Center S.L. Young Biomedical Research Center Rm. 466 Shipping address: 975 NE 10th Street, BRC 466 Oklahoma City, OK 73104-5419 Letter address: P.O. Box 26901, BRC 466 Oklahoma City, OK 73190 office: (405) 271-8300 lab: (405) 271-8313 fax: (405) 271-3910 e-mail: blaine-moo...@ouhsc.edu Faculty webpage: http://www.oumedicine.com/department-of-biochemistry-and-molecular-biology/faculty/blaine-mooers-ph-d- [1] X-ray lab webpage: http://www.oumedicine.com/department-of-biochemistry-and-molecular-biology/department-facilities/macromolecular-crystallography-laboratory [2] Small Angle Scattering webpage: http://www.oumedicine.com/docs/default-source/ad-biochemistry-workfiles/small-angle-scattering-links.html?sfvrsn=0 [3] From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] On Behalf Of Katherine Sippel [katherine.sip...@gmail.com] Sent: Tuesday, February 18, 2014 12:08 PM To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] High Salt Cryo Hi all, I'm looking for a cryo condition for high NaCl (3+ M) crystallization condition. I would do it the proper way, but our beam/cryostream is down. I've tried a bunch of things at the moment. Ethylene glycol and PEG 400 nuke the crystals immediately even at low concentrations. Prolonged exposure to glycerol and sucrose starts to break them down so I'm thinking that the diffraction will probably suffer. I can't find any reports of NaCl's viability as a cryosalt. I've got Paratone/Paraffin oil/Mitegen's LV cryo oil on tap but I was hoping to not put all my eggs in one basket. I tried the ISRDB database through archive.comhttps://urldefense.proofpoint.com/v1/url?u=http://archive.comk=7DHVT22D9IhC0F3WohFMBA%3D%3D%0Ar=ftLbjJYpc5s5JQz9Q6qd7uT7FxPLb4V0aIwH4RJhyZU%3D%0Am=Vjr4m%2Fds%2FdLGOVQoQ0x8PApF%2FzyGkSwsbIoq92CSnOk%3D%0As=3cfbf18821b5b59934971bf583cf3dd6e2ded91923c614e670857e10916c687e [4] without any luck (no search function). I've gone to the PDB searching for similar crystallization conditions and looked up the papers for their cryos, but they are all glycerol. Google gives me the same. I thought I'd see if anyone on the bb has an anecdotal this worked for us story. I would love to hear it. Thank you for your time, Katherine -- Nil illegitimo carborundum - Didactylos Links: -- [1] http://www.oumedicine.com/department-of-biochemistry-and-molecular-biology/faculty/blaine-mooers-ph-d- [2] http://www.oumedicine.com/department-of-biochemistry-and-molecular-biology/department-facilities/macromolecular-crystallography-laboratory [3] http://www.oumedicine.com/docs/default-source/ad-biochemistry-workfiles/small-angle-scattering-links.html?sfvrsn=0 [4]
Re: [ccp4bb] I/sigmaI or I/sigmaI
Dear Folmer Fredslund, Thanks for your help. Actually, I can only find the archives until APR-30-2003 at the archives. Where is the DEC-1-2003 archives? Regards, Qixu Cai 发件人: Folmer Fredslund folm...@gmail.com 日期: 2014年2月19日 星期三 下午10:09 至: Cai Qixu caiq...@gmail.com 抄送: CCP4BB@jiscmail.ac.uk CCP4BB@jiscmail.ac.uk 主题: Re: [ccp4bb] I/sigmaI or I/sigmaI Dear Qixu Cai, You can find information about where to find the archives here: http://www.ccp4.ac.uk/ccp4bb.php#archives Best regards, Folmer 2014-02-19 14:44 GMT+01:00 Cai Qixu caiq...@gmail.com: Dear Richard Gillilan, Where to find the archives of Dec 2003? I can only find the archives until 2007 at jiscmail. Thanks. Regards, Qixu Cai 发件人: Richard Gillilan r...@cornell.edu 答复: Richard Gillilan r...@cornell.edu 日期: 2014年2月16日 星期日 上午12:19 至: CCP4BB@JISCMAIL.AC.UK 主题: Re: [ccp4bb] I/sigmaI or I/sigmaI There was an informative discussion on this very topic back in Dec 1-2, 2003 if you browse the CCP4BB archives. Richard Gillilan MacCHESS On Feb 12, 2014, at 6:43 AM, Cai Qixu wrote: Dear all, Does the I/sigmaI in “Table 1” mean for I/sigmaI or I/sigmaI ? Thanks for your answer. Best wishes, Qixu Cai -- Folmer Fredslund
[ccp4bb] Postdoctoral research associate position in structural biology at the University of LIverpool
THE UNIVERSITY OF LIVERPOOL FACULTY OF HEALTH AND LIFE SCIENCES INSTITUTE OF INTEGRATIVE BIOLOGY DEPARTMENT OF BIOCHEMISTRY POSTDOCTORAL RESEARCH ASSOCIATE £31,644 - £36,661 pa An exciting opportunity has emerged to join the Molecular Biophysics Group to work on a BBSRC-supported project on structure-function-mechanism studies of Cu-nitrite reductase and membrane bound nitric oxide (NO) reductase and the role of metabolon complex formation in controlling levels of cytotoxic NO. The project will link crystallographic studies with techniques to probe factors that control delivery of electrons and protons to the active site of these enzymes. You will be responsible for using established molecular biology procedures underpinning the overexpression and purification of a number of enzymes and their mutants including membrane proteins and undertake crystallographic, spectroscopic and mechanistic aspects of the programme. You will have a chance to spend up to 4 weeks in the RIKEN laboratories in Japan as part of a collaborative programme alongside two PhD students working on membrane-bound nitric oxide reductases. You should have a PhD in a relevant area, with experience in protein crystallography and practical knowledge of molecular biology. Experience in membrane proteins is desirable. Excellent verbal and written communication skills are essential. The post is available for 3 years initially. Job Ref: R-584998 Closing Date: 30 March 2014 For full details, or to request an application pack, visit www.liv.ac.uk/working/job_vacancies/http://www.liv.ac.uk/working/job_vacancies/ or e-mail j...@liv.ac.ukmailto:j...@liv.ac.uk, please quote Job Ref in all enquiries Dr. Svetlana Antonyuk Senior Lecturer Institute of Integrative Biology University of Liverpool Biosciences Building Liverpool L69 7ZB -- Phone: +44(0)151-795 0349 Email:s.anton...@liverpool.ac.ukmailto:s.anton...@liverpool.ac.uk
[ccp4bb] New position
Dear colleagues, We have an opening for a structural biologist here in Frederick, MD. If not interested, please forward this email to those who might be. Thank you. Best regards, Bill http://jobs.leidos.com/job/Frederick-Lead-Structural-Biology-603181-%28NCI%29-Job-MD-21701/44050800/ http://jobs.leidos.com/job/Frederick-Lead-Structural-Biology-603181-%28NCI%29-Job-MD-21701/44050800/ __ Bill Gillette, Ph.D. (Contractor) Sr. Scientist, Protein Expression Laboratory Cancer Research Technology Program Leidos Biomedical Research, Inc. Frederick National Laboratory for Cancer Research (FNL) P.O. Box B, Frederick, MD 21702 Phone: 301-846-1029 william.gille...@nih.govgillet...@mail.nih.gov Notice: This communication may contain privileged or other confidential information. If you are not the intended recipient, or believe that you have received this communication in error, please do not print, copy, retransmit, disseminate or otherwise use the information. Please indicate to the sender that you have received this email in error and delete the copy you received.
[ccp4bb] Can not see density map when I turn off normalization in PYMOL
Hello there, I am making a fo-fc map for one ligand using pymol. I strictly followed the pymol wiki protocol (Display CCP4 Maps). Finally, I can get the ligand map using command: isomesh fo-fc_ligand, omitmap, 3, ligand, carve=2. However, the problem is the map I got from pymol is smaller than the one I can see in coot at the same contour level (3.0). So I gave a second trial based on the assumption that it may be caused by the mis-normalization. I input the command: unset normalize_ccp4_maps to stop PyMOL from normalizing a cpp4 map. After that I loaded my ccp4 map file and tried to do the same things as what I did for the first time. But I could not see any mesh net (density map) shown up. I check the command window. *PyMOLunset normalize_ccp4_maps* * Setting: normalize_ccp4_maps set to off.* * ObjectMapCCP4: Map Size 134 x 128 x 122* * ObjectMapCCP4: Map will not be normalized.* * ObjectMapCCP4: Current mean = -0.66 and stdev = 0.074981.* * ObjectMap: Map read. Range: -0.511 to 0.616* * Crystal: Unit Cell 200 300 100* * Crystal: Alpha Beta Gamma90.000 100.354 90.000* * Crystal: RealToFrac Matrix* * Crystal:0.0060 -0.0.0011* * Crystal:0.0.0045 -0.* * Crystal:0.0.0.0053* * Crystal: FracToReal Matrix* * Crystal: 2000. -34.5817* * Crystal:0. 3000.* * Crystal:0.0. 100* * Crystal: Unit Cell Volume 6993536.* * ExecutiveLoad: E:/ bdligand002.ccp4 loaded as bdligand002, through state 1.* * PyMOLisomesh fo-fc_ligand, bdligand002, 3, ligand, carve=2* * Executive: object fo-fc_ligand created.* * Isomesh: created fo-fc_ligand, setting level to 2* * ObjectMesh: updating fo-fc_ligand.* It seems like no error, but my ligand map, fo-fc_ligand has no density map shown up. I also tried to show the whole mesh at level 2.0 for bdligand002. I still could not see the density map. My pymol is version 1.3 in windows 8 operation system. Any help will be greatly appreciated! Thanks in advance hongshi
Re: [ccp4bb] Can not see density map when I turn off normalization in PYMOL
Dear Hongshi - I cant help but to first suggest that as this is the CCP4bb, the first thing you should try is using CCP4mg, if you want to make pretty pictures. If you just want to look at the map and work with it, you should use Coot. If still for some reason you want to use Pymol, you should ask the experts of Pymol, and its best to mail pymol-us...@lists.sourceforge.net These said, you do not specify what you really want to do. Is the ligand already there in the coordinates file? Then an fo-fc (difference) map should show you nothing, as the ligand is there and there should be no difference. If the ligands is not there in the coordinates file, and you see no density, then the ligand is simply not there. Hope these help, Tassos On 19 Feb 2014, at 18:30, hongshi WANG wrote: Hello there, I am making a fo-fc map for one ligand using pymol. I strictly followed the pymol wiki protocol (Display CCP4 Maps). Finally, I can get the ligand map using command: isomesh fo-fc_ligand, omitmap, 3, ligand, carve=2. However, the problem is the map I got from pymol is smaller than the one I can see in coot at the same contour level (3.0). So I gave a second trial based on the assumption that it may be caused by the mis-normalization. I input the command: “unset normalize_ccp4_maps” to stop PyMOL from normalizing a cpp4 map. After that I loaded my ccp4 map file and tried to do the same things as what I did for the first time. But I could not see any mesh net (density map) shown up. I check the command window. PyMOLunset normalize_ccp4_maps Setting: normalize_ccp4_maps set to off. ObjectMapCCP4: Map Size 134 x 128 x 122 ObjectMapCCP4: Map will not be normalized. ObjectMapCCP4: Current mean = -0.66 and stdev = 0.074981. ObjectMap: Map read. Range: -0.511 to 0.616 Crystal: Unit Cell 200 300 100 Crystal: Alpha Beta Gamma90.000 100.354 90.000 Crystal: RealToFrac Matrix Crystal:0.0060 -0.0.0011 Crystal:0.0.0045 -0. Crystal:0.0.0.0053 Crystal: FracToReal Matrix Crystal: 2000. -34.5817 Crystal:0. 3000. Crystal:0.0. 100 Crystal: Unit Cell Volume 6993536. ExecutiveLoad: E:/ bdligand002.ccp4 loaded as bdligand002, through state 1. PyMOLisomesh fo-fc_ligand, bdligand002, 3, ligand, carve=2 Executive: object fo-fc_ligand created. Isomesh: created fo-fc_ligand, setting level to 2 ObjectMesh: updating fo-fc_ligand. It seems like no error, but my ligand map, fo-fc_ligand has no density map shown up. I also tried to show the whole mesh at level 2.0 for bdligand002. I still could not see the density map. My pymol is version 1.3 in windows 8 operation system. Any help will be greatly appreciated! Thanks in advance hongshi
Re: [ccp4bb] Can not see density map when I turn off normalization in PYMOL
When you don't normalize the map you have to specify your contour level in whatever units the map came in. Your output says the stdev is 0.075 so I guess you need to contour at 0.225 to see the equivalent image. Dale Tronrud P.S. I feel compelled to note that what the program is reporting as the standard deviation is really the root mean square deviation from zero. The standard deviation of a map is a much more subtle quantity as discussed recently in PNAS. On 02/19/2014 09:30 AM, hongshi WANG wrote: Hello there, I am making a fo-fc map for one ligand using pymol. I strictly followed the pymol wiki protocol (Display CCP4 Maps). Finally, I can get the ligand map using command: isomesh fo-fc_ligand, omitmap, 3, ligand, carve=2. However, the problem is the map I got from pymol is smaller than the one I can see in coot at the same contour level (3.0). So I gave a second trial based on the assumption that it may be caused by the mis-normalization. I input the command: “unset normalize_ccp4_maps” to stop PyMOL from normalizing a cpp4 map. After that I loaded my ccp4 map file and tried to do the same things as what I did for the first time. But I could not see any mesh net (density map) shown up. I check the command window. /PyMOLunset normalize_ccp4_maps/ / Setting: normalize_ccp4_maps set to off./ / ObjectMapCCP4: Map Size 134 x 128 x 122/ / ObjectMapCCP4: Map will not be normalized./ / ObjectMapCCP4: Current mean = -0.66 and stdev = 0.074981./ / ObjectMap: Map read. Range: -0.511 to 0.616/ / Crystal: Unit Cell 200 300 100/ / Crystal: Alpha Beta Gamma90.000 100.354 90.000/ / Crystal: RealToFrac Matrix/ / Crystal:0.0060 -0.0.0011/ / Crystal:0.0.0045 -0./ / Crystal:0.0.0.0053/ / Crystal: FracToReal Matrix/ / Crystal: 2000. -34.5817/ / Crystal:0. 3000./ / Crystal:0.0. 100/ / Crystal: Unit Cell Volume 6993536./ / ExecutiveLoad: E:/ bdligand002.ccp4 loaded as bdligand002, through state 1./ / PyMOLisomesh fo-fc_ligand, bdligand002, 3, ligand, carve=2/ / Executive: object fo-fc_ligand created./ / Isomesh: created fo-fc_ligand, setting level to 2/ / ObjectMesh: updating fo-fc_ligand./ It seems like no error, but my ligand map, fo-fc_ligand has no density map shown up. I also tried to show the whole mesh at level 2.0 for bdligand002. I still could not see the density map. My pymol is version 1.3 in windows 8 operation system. Any help will be greatly appreciated! Thanks in advance hongshi
Re: [ccp4bb] Can not see density map when I turn off normalization in PYMOL
Hi Hongshi, I think Dale is right. When you turn off the normalization, the level value you need to put in the isomesh command should be the absolute electron density value of the map instead of how many RMSD. In your command window log, this line shows the range of the density values in your map: ObjectMap: Map read. Range: -0.511 to 0.616 As you can see the highest value is 0.616, no wonder you see nothing when you tell the program to cut off at 3.0. And the line above it shows the RMSD, marked as stdev: ObjectMapCCP4: Current mean = -0.66 and stdev = 0.074981. So for RMSD=3.0, the density cut off of this map is 0.075x3.0=0.225. Your command then is: isomesh fo-fc_ligand, bdligand002, 0.225 Or you can look at the coot window to find out what the density cut off is being used (map level=x. e/A^3 ), if you are not too sure about the RMSD calculation. An additional note: the same applies to UCSF Chimera, where the volume data (maps) control also requires you to specify the cut off or gradient in real map values instead of RMSD. Zhijie From: hongshi WANG Sent: Wednesday, February 19, 2014 3:47 PM To: Zhijie Li Subject: Re: [ccp4bb] Can not see density map when I turn off normalization in PYMOL Hi Zhijie, Thanks for your help. Once i turned off the normalization in pymol. I could not see any density map. hongshi On Wed, Feb 19, 2014 at 11:21 AM, Zhijie Li zhijie...@utoronto.ca wrote: Hi Hongshi, If you just put isomesh fo-fc_ligand, omitmap, 3 do you seem anything in the unit cell? If you do then please check if around your ligand there is any density. Zhijie From: hongshi WANG Sent: Wednesday, February 19, 2014 12:30 PM To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] Can not see density map when I turn off normalization in PYMOL Hello there, I am making a fo-fc map for one ligand using pymol. I strictly followed the pymol wiki protocol (Display CCP4 Maps). Finally, I can get the ligand map using command: isomesh fo-fc_ligand, omitmap, 3, ligand, carve=2. However, the problem is the map I got from pymol is smaller than the one I can see in coot at the same contour level (3.0). So I gave a second trial based on the assumption that it may be caused by the mis-normalization. I input the command: “unset normalize_ccp4_maps” to stop PyMOL from normalizing a cpp4 map. After that I loaded my ccp4 map file and tried to do the same things as what I did for the first time. But I could not see any mesh net (density map) shown up. I check the command window. PyMOLunset normalize_ccp4_maps Setting: normalize_ccp4_maps set to off. ObjectMapCCP4: Map Size 134 x 128 x 122 ObjectMapCCP4: Map will not be normalized. ObjectMapCCP4: Current mean = -0.66 and stdev = 0.074981. ObjectMap: Map read. Range: -0.511 to 0.616 Crystal: Unit Cell 200 300 100 Crystal: Alpha Beta Gamma90.000 100.354 90.000 Crystal: RealToFrac Matrix Crystal:0.0060 -0.0.0011 Crystal:0.0.0045 -0. Crystal:0.0.0.0053 Crystal: FracToReal Matrix Crystal: 2000. -34.5817 Crystal:0. 3000. Crystal:0.0. 100 Crystal: Unit Cell Volume 6993536. ExecutiveLoad: E:/ bdligand002.ccp4 loaded as bdligand002, through state 1. PyMOLisomesh fo-fc_ligand, bdligand002, 3, ligand, carve=2 Executive: object fo-fc_ligand created. Isomesh: created fo-fc_ligand, setting level to 2 ObjectMesh: updating fo-fc_ligand. It seems like no error, but my ligand map, fo-fc_ligand has no density map shown up. I also tried to show the whole mesh at level 2.0 for bdligand002. I still could not see the density map. My pymol is version 1.3 in windows 8 operation system. Any help will be greatly appreciated! Thanks in advance hongshi
Re: [ccp4bb] Symmetry problem
Monika, There are several possible causes for the problem you are encountering, but your description is a little too vague to discern them. Scenario 1) You ran phaser with the option all possible spacegroups, for several different components of your crystal, setup individually, and the runs do not agree on the best spacegroup? -- In that case, phaser had problems determining the correct spacegroup, I'd suggest you search for all components, but in separate runs for each possible spacegroup. Scenario 2) You assumed your spacegroup assignment was correct, and ran MR for each of your components individually, and when you display the solutions, they overlap. In this case, you might have your solutions on different origins. The best way out is to use the first solution as a fixed solution, which is possible in most MR programs, and then search for the next component. There might be other scenarios, if you describe your situation in more detail (how many components in the crystal setup, what program you used, how you used it, and what you mean by different symmetries), we might be able to help you better, Cheers, Jens On Tue, 2014-02-18 at 14:59 -0300, Monika Coronado wrote: Dear, Does anyone know how to merge two molecules with different symmetry? I will explain: I have done the molecular replacement using the domains of the molecules separately, now I have to put all together, however they have a different symmetry. I will appreciate any kind of help. Regards, Mônika -- __o _`\,_ (*)/ (*) -+-+-+-+-+-+- * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * ...E tudo muda... * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * * *
[ccp4bb] keep ligand conformation
Dear all, I tried to keep a conformation of a ligand during refinement with Refmac5 (Ver. 5.7.0032), and I put a harmonic restraint as, external harmonic residues from 600 A to 600 A 600 is the residue number of the ligand. I checked structures with and without the harmonic restraint, but no change was found between the two structures at all. I also tried with and without sigma weight, but nothing changed again. external harmonic restraints worked for protein parts. I would appreciate any suggestions and comments. Thank you so much in advance, Koji
