Re: [ccp4bb] Water molecules after refinement
This may be a useful paper to read, although it is a bit dated: Acta Cryst. (1999). D55, 479-483 How many water molecules can be detected by protein crystallography? In general, 3 A resolution is beyond where you can reliably add waters (I start adding them only at 2.7-2.8 A resolution or better). If you do want to add waters, make sure they make sense (H-bonding/distance) and do not have positive Fo-Fc density after refinement, in which case they could be metal ions. If after refinement the (2Fo-Fc) density disappears then whatever you thought was there isn't. best, bert From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] on behalf of jeorgemarley thomas [kirtswab...@gmail.com] Sent: Wednesday, November 19, 2014 6:33 AM To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] Water molecules after refinement Dear All, I am sorry to ask this simple question, but I really need suggestions for this. As the refinement has been done at 3.0 Angstrom after refinement the water molecules were added by using "find waters" in coot. After adding water refinement was done using Refmac 05. Now when I look on the added water molecules some has density around it and some does not (even no red density around it). Should I remove later water molecules. and again do the refinement ? Please give some explanation also for this. as the R free and R meas has reduced to some extent, also the r m s d contour of map level was kept at 1.09 after refinement. I welcome your suggestions, thanks in advance. Regards Jeorge
Re: [ccp4bb] superpose bug?
Correct syntax is superpose A.pdb B.pdb -o C.pdb Doing superpose A.pdb B.pdb C.pdb will result in *multiple* structure alignment of all three structures (i.e. C.pdb ought to be an input, rather than output, file here). In general, superpose XYZ1.pdb XYZ2.pdb XYZ3.pdb . XYZN.pdb will do multiple aligment of all N structures. Run superpose without parameters in order to get description of all command-prompt options. Hope this helps, Eugene From: Lijun Liu [lijunli...@gmail.com] Sent: Wednesday, November 19, 2014 12:17 AM To: ccp4bb Subject: [ccp4bb] superpose bug? Hi: I have a bunch of pdb files and would like to superpose them to one of them based on secondary structure matching. I would like to get a script to do so (not a one-by-one job by hand). I tried superpose but it seemed to have some problems with output option. Assuming C.pdb is the output pdb, and A.pdb and B.pdb are inputs. superpose A.pdb B.pdb (this works, without output) superpose A.pdb B.pdb C.pdb (* ERROR #15 READ:) Adding -s option get same errors (without output pdb it works). When running within ccp4i with output C.pdb, it does work. I tried to generate an input script from ccp4i running. Unfortunately, running with the particular "superpose using S-S-M" option did not generate an input card (other options OK, what a bad luck). I appreciate a working script (ccp46.4.0) or a "debugged" source code for compilation. Thanks. Lijun Liu -- Scanned by iCritical.
Re: [ccp4bb] Fwd: [ccp4bb] B-factor blurring
Dear Mike, I am not sure what you mean by 'reference'. When you multiply your data with exp(-Bs^2) with B > 0 you weight your reflections depending on resolution so that high resolution data (probably including noise) are supressed while low resolution data a underlined. Using the term 'B' with its relationship to the ADPs is probably only a way to make it sound more fancy. You can call this textbook knowledge, maybe that's why there is not a real reference? Best, Tim On 11/19/2014 01:34 AM, Mike Lawrence wrote: > My sincere thanks to all who are responding to my request below. > > To be explicit, my question relates to B-factor blurring (+B correction), not > to B-factor sharpening (-B correction). > > thanks > > Mike > > > Begin forwarded message: > >> From: Mike Lawrence >> Subject: [ccp4bb] B-factor blurring >> Date: 18 November 2014 12:01:07 pm AEDT >> To: CCP4BB@JISCMAIL.AC.UK >> Reply-To: Mike Lawrence >> >> Dear all >> >> can anyone help me with literatures references to B-factor blurring as a >> technique to reveal low resolution features in an electron density map? I >> have seen the poster from Andrea Thorn at >> >> http://shelx.uni-ac.gwdg.de/~athorn/pdf/thorn_iucr2014_poster.pdf >> >> but was wondering if there were any alternative references? >> >> with many thanks! >> >> Mike >> >> > > > > __ > The information in this email is confidential and intended solely for the > addressee. > You must not disclose, forward, print or use it without the permission of the > sender. > __ > -- Dr Tim Gruene Institut fuer anorganische Chemie Tammannstr. 4 D-37077 Goettingen GPG Key ID = A46BEE1A signature.asc Description: OpenPGP digital signature
Re: [ccp4bb] Water molecules after refinement
Dear Jeorge, as you add water molecules to a model, the R _and_ Rfree are bound to drop. It is important to do it the other way round: let coot add waters, then check the added waters in coot if they make chemically sense, then run refmac5. Don't trust automatic procedures more than your chemical education, no matter how fancy. At 3A I would be careful adding any waters at all, you might just fit noise. Best, Tim On 11/19/2014 07:33 AM, jeorgemarley thomas wrote: > Dear All, > > I am sorry to ask this simple question, but I really need suggestions for > this. As the refinement has been done at 3.0 Angstrom after refinement the > water molecules were added by using "find waters" in coot. After adding > water refinement was done using Refmac 05. Now when I look on the added > water molecules some has density around it and some does not (even no red > density around it). Should I remove later water molecules. and again do the > refinement ? Please give some explanation also for this. as the R free and > R meas has reduced to some extent, also the r m s d contour of map level > was kept at 1.09 after refinement. I welcome your suggestions, thanks in > advance. > > > Regards > > Jeorge > -- Dr Tim Gruene Institut fuer anorganische Chemie Tammannstr. 4 D-37077 Goettingen GPG Key ID = A46BEE1A signature.asc Description: OpenPGP digital signature
Re: [ccp4bb] superpose bug?
Hi Eugene, would you update the documentation? A canonical 'man superpose' produces superpose foo_1st.pdb [-s CID1] foo_2nd.pdb [-s CID2] [ foo_out.pdb ] where [-s CID1/2] are optional selection strings in [1]MMDB convention, and [foo_out.pdb] is an optional output file. i.e. the syntax Lijun Liu used. Cheers, Tim On 11/19/2014 10:33 AM, Eugene Krissinel wrote: > Correct syntax is > > superpose A.pdb B.pdb -o C.pdb > > Doing > > superpose A.pdb B.pdb C.pdb > > will result in *multiple* structure alignment of all three structures (i.e. > C.pdb ought to be an input, rather than output, file here). In general, > > superpose XYZ1.pdb XYZ2.pdb XYZ3.pdb . XYZN.pdb > > will do multiple aligment of all N structures. > > Run superpose without parameters in order to get description of all > command-prompt options. > > Hope this helps, > > Eugene > > > > From: Lijun Liu [lijunli...@gmail.com] > Sent: Wednesday, November 19, 2014 12:17 AM > To: ccp4bb > Subject: [ccp4bb] superpose bug? > > Hi: > > I have a bunch of pdb files and would like to superpose them to one of > them based on secondary structure matching. I would like to get a > script to do so (not a one-by-one job by hand). > > I tried superpose but it seemed to have some problems with output > option. Assuming C.pdb is the output pdb, and A.pdb and B.pdb are > inputs. > > > superpose A.pdb B.pdb (this works, without output) > superpose A.pdb B.pdb C.pdb (* ERROR #15 READ:) > Adding -s option get same errors (without output pdb it works). > > When running within ccp4i with output C.pdb, it does work. > > > I tried to generate an input script from ccp4i running. Unfortunately, > running with the particular "superpose using S-S-M" option did not > generate an input card (other options OK, what a bad luck). > > I appreciate a working script (ccp46.4.0) or a "debugged" source code > for compilation. Thanks. > > Lijun Liu > -- Dr Tim Gruene Institut fuer anorganische Chemie Tammannstr. 4 D-37077 Goettingen GPG Key ID = A46BEE1A signature.asc Description: OpenPGP digital signature
Re: [ccp4bb] superpose bug?
Yes, certainly, guilty as sin :) Thanks, Eugene From: Tim Gruene [t...@shelx.uni-ac.gwdg.de] Sent: Wednesday, November 19, 2014 11:22 AM To: Krissinel, Eugene (STFC,RAL,SC); ccp4bb Subject: Re: [ccp4bb] superpose bug? Hi Eugene, would you update the documentation? A canonical 'man superpose' produces superpose foo_1st.pdb [-s CID1] foo_2nd.pdb [-s CID2] [ foo_out.pdb ] where [-s CID1/2] are optional selection strings in [1]MMDB convention, and [foo_out.pdb] is an optional output file. i.e. the syntax Lijun Liu used. Cheers, Tim On 11/19/2014 10:33 AM, Eugene Krissinel wrote: > Correct syntax is > > superpose A.pdb B.pdb -o C.pdb > > Doing > > superpose A.pdb B.pdb C.pdb > > will result in *multiple* structure alignment of all three structures (i.e. > C.pdb ought to be an input, rather than output, file here). In general, > > superpose XYZ1.pdb XYZ2.pdb XYZ3.pdb . XYZN.pdb > > will do multiple aligment of all N structures. > > Run superpose without parameters in order to get description of all > command-prompt options. > > Hope this helps, > > Eugene > > > > From: Lijun Liu [lijunli...@gmail.com] > Sent: Wednesday, November 19, 2014 12:17 AM > To: ccp4bb > Subject: [ccp4bb] superpose bug? > > Hi: > > I have a bunch of pdb files and would like to superpose them to one of > them based on secondary structure matching. I would like to get a > script to do so (not a one-by-one job by hand). > > I tried superpose but it seemed to have some problems with output > option. Assuming C.pdb is the output pdb, and A.pdb and B.pdb are > inputs. > > > superpose A.pdb B.pdb (this works, without output) > superpose A.pdb B.pdb C.pdb (* ERROR #15 READ:) > Adding -s option get same errors (without output pdb it works). > > When running within ccp4i with output C.pdb, it does work. > > > I tried to generate an input script from ccp4i running. Unfortunately, > running with the particular "superpose using S-S-M" option did not > generate an input card (other options OK, what a bad luck). > > I appreciate a working script (ccp46.4.0) or a "debugged" source code > for compilation. Thanks. > > Lijun Liu > -- Dr Tim Gruene Institut fuer anorganische Chemie Tammannstr. 4 D-37077 Goettingen GPG Key ID = A46BEE1A -- Scanned by iCritical.
[ccp4bb] Structural dynamics in cellular communication
Dear all, I would like to attract your attention to a conference entitled 'Structural dynamics in cellular communication', organised by the Flemish Institute for Biotechnology (VIB) in Brussels, February 9-10, 2014. More information can be found on the events website: http://www.vibconferences.be/event/structural-dynamics-in-cellular-communication Register before the end of November and save 20% on the registration price (now €220 instead of €275). This registration fee covers your entrance, attendance to all talks, lunches and coffee breaks and the first day's closing reception. Deadline for abstract submission is November 30, 2014. Kind regards, Inge Dr. Inge Van Molle VIB Structural Biology Research Center Oxidative Stress Signalling Structural Biology Brussels Vrije Universiteit Brussel Building E, room 4.16, Pleinlaan 2, 1050 Brussel, Belgium Tel. 003226291992 Mobile 0032486521278
Re: [ccp4bb] how to combine two derivative datasets that are not isomorphous?
How non-isomorhous is non-isomorphous? Can you give the cells and putative Spacegroups for A a B & b? You can often get information at low resolution for this sort of problem if there is some level of similarity. Eleanor On 18 November 2014 21:15, David Schuller wrote: > You don't mention any quality indicators on your derivatives, nor > resolution. > > Presuming they actually have some decent phasing power, you may be able to > generate phases & maps using SIR phasing + solvent flattening. If you can > do that for each isomorphous set, then you could combine them using > multiple space group noncrystallographic averaging. > > Or, if the derivatives contain atoms with anomalous scattering, you could > solve a derivative alone with SAD methods, or a derivative + native set > with SIRAS. Once you get crude maps, you can once again try to combine the > two sets with multiple space group averaging. > > As for MR, there are lots of things you can try. Once again, you have > provided no detail on what the sequence similarity is, or any other factor > that would allow us to judge the likelihood of success. > You could search again with the model clipped down to poly-Ala. You could > search again with any external loops trimmed off. If it is a multiple > domain molecule, you could search individually with single domains. You > could find additional search models, and search with a suite rather than > single model. MR programs will suck up as much free time as you can provide > them. > > > > On 11/18/14 15:01, joy yang wrote: > > Hi All, > > I have two derivative datasets (heavy atom A and B) and two native > datasets (a and b), A and a are isomorphous, B and b are isomorphous, > however, a and b (or A and B) are not isomorphous. > > I was able to make two difference patterson maps (FA-Fa and FB-Fb) and > search for heavy atoms against them, the possible positions of heavy atom A > and B are very close to each other in the unit cell (which seems to me that > I am very close to a right phase, though not there yet), I am wondering if > there is any means for me to combine the information from FA-Fa and FB-Fb > as the two native datasets are not isomorphous? And also, I have a homology > model which I tried molecular replacement and failed, is there any means > for me to combine the information from the model too? > > Best, > > Bei > > > > -- > === > All Things Serve the Beam > === >David J. Schuller >modern man in a post-modern world >MacCHESS, Cornell University >schul...@cornell.edu > >
[ccp4bb] Postdoctoral positions available at the University of Pittsburgh, Pittsburgh, PA, United States - Structural and molecular pharmacology of GPCRs
We have immediate openings for postdoctoral fellows in the lab of Dr. Cheng Zhang in the Department of Pharmacology and Chemical Biology at School of Medicine, University of Pittsburgh at Pittsburgh, PA, United States. Our lab is a newly established lab that focus on the studies of a family of cell surface receptors - G protein-coupled receptors (GPCRs). The aim is to understand the molecular mechanisms of the signal transduction by GPCRs, which represent 30-40% of current drug targets, and to explore the molecular pharmacology to facilitate future drug design. The initial research effort will focus on the structural characterization of several GPCRs that function in inflammation and calcium metabolism in complex with different ligands and signaling effectors. The main approach is X-ray crystallography, complemented by EM and NMR methods. The obtained structural information will be used in the following structure-based drug design to find new chemicals as potential drugs with desired pharmacological properties. Also structure-based development of macro-biomolecules such as antibodies and peptides as potential pharmaceuticals that can modulate the functional properties of GPCRs is another research direction. So far several GPCRs as important drug targets have been successfully prepared in the lab, which are ready for further structural studies. The potential candidates are expected to perform structural studies on these receptors and/or develop other biophysical and cell-based experiments to examine the signaling behaviors of these receptors in respond to different ligands/drugs. Interested candidates should be highly motivated and have a PhD with strong background in one or several of the followings: 1) Molecular cloning and recombinant protein expression and purification. Experienced in protein characterization using various biochemical and biophysical methods such as pull-down, western blot, mass spec, ITC etc. 2) Protein X-ray Crystallography. 3) Recombinant protein expression in mammalian cells. Making stable cell lines. FACS setups. Experience with confocol or TIRF is optional but would be a plus. 4) Antibody production and engineering. Producing antibodies in hybridoma cells. Antibody characterization by ELISA. Experience in phage display or yeast display would be a major plus. We offer a quite exciting and vibrating working environment. There are several laboratories in the department that focus on the studies of GPCRs. You will be working with enthusiastic professionals studying GPCRs from difference perspectives. An competitive salary will be provided based on previous research experience and accomplishments. The initial appointment will be one year with the potential to renew every year depending on the progress of the projects. As for living, Pittsburgh is a very nice city to live. It has been elected as one of the most livable cities in the US for years with a diversity of people and culture. For more information about the lab and the department, please see: http://www.pharmacology.us/Faculty/ChengZhang To apply, please send a CV with cover letter to Dr. Cheng Zhang at chengzh1...@gmail.com. 2-3 reference letters will be requested if you are shortlisted for call interview.
