Re: [ccp4bb] molecular replacement with poor model

2014-12-12 Thread xaravich ivan
What is the resolution of your data? I have been able to get a solution for my protein with 30% identity but my resolution of data was 1.4 Angs. I believe to get a solution at 18% identity your search model has to be very close, like using Robetta to make the 3 mer and 9mer peptides and then work f

Re: [ccp4bb] molecular replacement with poor model

2014-12-12 Thread Randy Read
Dear Ursula, 18% identity is really in the twilight zone for molecular replacement, where it may work but there are certainly no guarantees. However, there’s a feature in Phaser that is useful for this kind of problem, i.e. the “rotate around” option, where you take advantage of knowing the ap

Re: [ccp4bb] asymmetric homotrimer in the asu

2014-12-12 Thread jtame
Dear Jose Your email expressed far more clearly than mine the problem of symmetry, and I had completely forgotten this discussion in the classic MWC paper. The point about measuring the molecular weight in solution is to find IF the protein forms an oligomer or not - AUC can give some idea of sha

Re: [ccp4bb] molecular replacement with poor model

2014-12-12 Thread Pavel Afonine
Hi Ursula, I also tried to just superimpose the complete model onto the partial > solution. This results in quite nice packing, but doesn't refine. Is there > a rigid program refinement program with very large convergence? > depending on what you call "very large", this may be helpful: Automat

[ccp4bb] molecular replacement with poor model

2014-12-12 Thread Ursula Schulze-Gahmen
I am trying molecular replacement with a very poor model. The model consists mainly of 1 long helix and two slightly bent antiparallel helices. After dividing it into 2 fragments, I was able to find a solution for one of the fragments ( at least I think so after looking at maps, packing, refinement

Re: [ccp4bb] asymmetric homotrimer in the asu

2014-12-12 Thread Kushol Gupta
Hi everyone, Just wanted to second the SAXS suggestion: the approach should provide a very rigorous way of testing the candidate models apparent in the crystallographic lattice, and can assist with questions of mass. (among the most common applications) With regards to the mass questi

Re: [ccp4bb] asymmetric homotrimer in the asu

2014-12-12 Thread David Briggs
Hi Hay, I think SAXS should be more than capable of discriminating between a 12.5 kDa monomer vs ~37.5 kDa trimer. Lysozyme is a useful standard used in SAXS (as with most structural biology!), and Lysozyme is only slightly larger than your proteins. Cheers, Dave On Fri Dec 12 2014 at 5:13:26

Re: [ccp4bb] asymmetric homotrimer in the asu

2014-12-12 Thread Hay Dvir
Tanner: Thanks, GREAT reference on asymmetric homo oligomers! SAXS sounds like a good idea for a bit larger particles. I'm afraid it might be very difficult to get enough resolution to resolve oligomerization of a rather small 12.5 kDa protein like ours, but will look into it more closely. Joes

Re: [ccp4bb] asymmetric homotrimer in the asu

2014-12-12 Thread Tanner, John J.
Two thoughts on asymmetric oligomers. 1. Here is a recent survey of asymmetric homodimers in the PDB. I know you are looking for trimers, but at least this provides a precedent for asymmetric oligomers. Swapna LS, Srikeerthana K, Srinivasan N. Extent of structural asymmetry in homodimeric pro

[ccp4bb] Beamline scientist positions, EMBL@PETRA3

2014-12-12 Thread Thomas R. Schneider
Dear all, PETRA III will restart next spring and we are looking for people to join the EMBL team operating and developing the P13/P14 beamlines and related facilities. The positions available comprise user support and the possibility to carry out research projects. EMBL and the DESY Campus offe

Re: [ccp4bb] asymmetric homotrimer in the asu

2014-12-12 Thread Jose Manuel Duarte
Dear Hay Your post prompted me to respond, since I think the issue of symmetry is extremely important. I would like to reinstate here what should be obvious to everyone: a stable asymmetric assembly of proteins in solution is essentially impossible (or at most very very unlikely), purely bec

Re: [ccp4bb] asymmetric homotrimer in the asu

2014-12-12 Thread Hay Dvir
Dear Jeremy, Indeed, we also incline to think of it as a monomer in solution, but still quite un-eased by the extensive interactions in the asu being merely as a result of a crystallization artifact. As you said, we may need to rely more heavily on biochemical analysis and since SEC wasn't clea