Dear experts, here is a Refmac question (or at least a refinement question):
the case: data set was collected at the synchrotron with high intensity or better high total dose. Data set scales to 1.2A resolution. Refinement with refmac5_0066, using hydrogens, anisotropic B-factors, automatic weight (actually 17.9), 235 amino acids, approx. 450 waters, several ions and buffer/crystallization compounds, a hell of second and third side chain conformations, present R-fac 12.6, free-R 15.6. I think I'm seeing a (partial) backbone break (negative DelFwt density for backbone atoms at 3.5sig within coot) due to radiation damage in a surface loop. I see clear missing COOH- and Guanidinium-groups in other parts of the model!! As a consequence, the protein adopts a second (backbone) conformation (clear positive DelFwt at 4.0sig) for an alpha-helix. But some of the second conformation residues are not visible. Summary: conformation A (aa206-219) ~ 70% occupancy, conformation B (aa212-219) ~ 30% occupancy, no density (flexibility??? / deletion ???) for aa206-211. Refmac always link aa212, conformation B to aa211, conformation A. But, the CA distance between Calpha atoms of aa212 is / should be about 5A. How to address the "non-restraint" between these amino acids, or vice versa to address a (partial) chain break? Furthermore I see "partial" waters at a (second) side chain conformation/position. Is it also possible to release the restraints for these waters to atoms of the second side conformation? It is really important to do it right, since the second conformation residues change the active site with a bound ligand. Every hint or work-around is highly appreciated. May be I have to use SHELXL? Regards Michael -- Dr. Michael Weyand mail: michael.wey...@mpi-dortmund.mpg.de Max-Planck-Institut fuer molekulare Physiologie, Otto-Hahn-Strasse 11 D-44227 Dortmund, Germany, fon: +49(0)231-133-2738, fax: +49(0)231-133-2797
