Ethan Merritt schrieb:
Hi all,I have an interesting problem case at the moment. We have crystallized the same 2-domain protein in two different,i.e. non-isomorphous, crystal forms.There is a decent homologous structure for one domain (about 40% of the total), but no known homologous structure for the larger second domain. Phaser easily places the probe structure in both unit cells, but recovering the remaining 60% of the structure from that starting point is problematic. In both cells there is only a single copy of the protein per a.s.u., Resolution is on the order of 2.6A, and the solvent content is relatively low. So no help from NCS or solvent flattening for either structure by itself. Yes, we can start searching for derivatives or SeMet data, but meanwhile I am looking for advice on the current generation of tools available for cross-crystal density averaging. Are there any that tie in to auto-tracing in Arp/wArp or resolve? Is there any chance of eventually doing joint refining of both structures simultaneously?
Ethan, I would try toa) chop the homologous structure into 2ndary structure elements and rigid-body refine those, b) adjust the sequence and fit the mutated side chains into their density, using Coot
c) do some restrained refinement in refmaccheck the R_free after a), b), c) and pick the best structure based on that. Then run buccanneer - it has worked for me once in a similar case.
best, Kay -- Kay Diederichs http://strucbio.biologie.uni-konstanz.de email: kay.diederi...@uni-konstanz.de Tel +49 7531 88 4049 Fax 3183 Fachbereich Biologie, Universität Konstanz, Box M647, D-78457 Konstanz This e-mail is digitally signed. If your e-mail client does not have the necessary capabilities, just ignore the attached signature "smime.p7s".
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