Hi For your SAD / MAD data, firstly after merging can you see any peaks in the anomolous difference patterson map - this is critical. You could try using SOLVE / RESOLVE or SHARP / AUTOSHARP to identify sites and to calculate phases, as you have a little bit more control over the process. You don't mention you Space group - are there any ambiguities here? If you have an poor initial MR model, together with some poor anomolous phases then you could use SHARP for phased molecular replacement.
For PHASER we have found that (unsurprisingly) in low identity cases it can be very sensitive to the quality of the MR model - have you carefully pared down the model, removed loops, truncated non identical positions to C-beta etc? Its a little unclear from your mail (I may have missed some of this thread though) how many mols you think there are in the ASU - my advice is to start simple - ask PHASER to find one molecule (hit the all possible SG flag) and see what the LLG and Z-score is, if you get a significant hit, then fix and re-run asking it to find another (or just rerun asking it to find two) and so on - again LLG and Z-score are critical here. You may have to play with the clashing factor as sometimes "good" hits are rejected because of clashes between bits of the MR model that should have been deleted (if you consistently see logical packing but two bits of the structure clash that could be reasonably shifted then adjust your MR model accordingly) ! At each stage inspect the maps, the scores and the packing really carefully. If you have more than one template structures try an ensemble of models as well. Cheers J Yi Xue <[EMAIL PROTECTED]> wrote:> > So far, the native crystals diffracted best to 2.4A. The MAD data > diffracted to 2.6~2.7A. We attempted to use phenix.hyss to identify > copper > atoms, and the program had hard time to identify the sites. > > The protein: Cu ratio is around 1:1, which is decided by ICP-AES > measurement > of the crystallization sample, not derived from the number of peaks in > the > anomalous map. > > The crystal contains copper as a cofactor, not a soaking derivative > thing. > > As to oligomerization, it could be dimer or trimer, however, we don't > have a > model for it, since we dont know exactly, how does the drug link the > monomers. > > I used phaser to do the MR, basically, what I did was to search one copy > each time, the top 20 solutions will be used to start the search of the > next copy. It stuck after finding four copies, and, I tried to change the > some paramters for searching, such as percentage for peaks, it did not > help. > I would love the hear from experienced people about some tricks using > phaser > to solve some difficult MR cases. > > > thanks a lot > Yi -- Professor James Whisstock NHMRC Principal Research Fellow / Monash University Senior Logan fellow Department of Biochemistry and Molecular Biology Monash University, Clayton Campus, PO Box 13d, VIC, 3800, Australia +613 9905 3747 (Phone) +613 9905 4699 (Fax) +61 418 170 585 (Mobile)
