Dear all! I have two questions:
First - I am refining a structure containing a modified residue with a ligand bound. This ligand is present in two stereoisomers and it seems like we have both in the structure. When refining the structure with ALIG and BLIG (each with 0.5 occupancy) defined in the coordinate file, REFMAC5 moves the ligands apart. In the ligand there is an electron rich atom with a clearly defined position in the electron density map (observed at 9 sigma). Prior to refinement, the electron rich atom from the two stereoisomers are positioned in the centre of the strong density feature. After ten cycles of refinement the atoms are separated (by about 1 Å) and found in two different positions, neither of them coinciding with the strong density. I dont understand why the stereoisomers are moved apart. The REFMAC log file does not give any warnings about close contacts between the atoms, but still they are moved during refinement and I dont know how to avoid this! Any help is much appreciated! To contribute to the confusion, the second problem is actually quite the opposite I have a non-covalent ligand bound in two alternative, partly overlapping sites. The sites induce distinct structural changes of the protein. For example, a Trp is moved to sandwich the ligand in one of the sites whereas this conformation of the Trp is not consistent with the second binding site. Ive tired to refine this structure using REFMAC5 and naming the Trp ATRP/BTRP and the corresponding ligand ALIG/BLIG in the coordinate file. Since the ATRP density map is stronger, the BTRP moves towards this density, even though this is create clashes between the BTRP and the BLIG. The program doesnt seem to link the two sites of the ligand with the corresponding conformations of the Trp. Is it possible to link the ALIG to the ATRP conformation of the protein and ligand? Sincerely, Andreas
