Dear collegues, I'm working with a drug complexed protein structure that is having major twinning issues. The drug has a single Br atom on a benzene ring, which I'd like to use for orienting the drug in the binding site. I have various anomalous data sets, ranging from 3.0A resolution, all scaled into P222 with a Rlin of .125.
Using MR, the twin law (k, h, -l) and NCS restraints, I can confidently solve the structure without anomalous, and the drug density is clear in the Fo-Fc map, with Rw/Rf at ~.26/.29 and a space group of P21221. It might be important to note that any simulated annealing I've tried invariably increases the Rfree by 2-3%, so I've scraped it. As you can imagine, when using the twinned data, the anomalous maps are weak and random. I've used the Phenix "detwin" option in Xtriage to see if I can pull the anomalous signal out of it. If I use the .mtz file that is output for MR and calculate the anomalous maps, it looks promising. The twin fraction for the one particular dataset I've been using is estimated at approx .48. Is this too close to 50% to do the detwinning? Now I'm wondering how to properly refine this further. I'm assuming that since I've "detwinned" the data, I do refinement without the twin law. But that gives an initial Rf of .38 when using it gives .31. Since I've already solved the structure without using the anomalous flag, can I just use the "detwinned" reflections and the refined structure to calculate an anomalous map (without having to redo the refinement)? Mainly, my main question is about how to tease out and properly refine the anomalous data from a twinned structure. Also, how much of a difference will it make to scale into P222 versus P21212. And, if I have quite high redundancy, should I "scale anomalous" in HKL2000 or just use the "anomalous" flag? Any help on refining this twinned structure would be greatly appreciated! Thanks, Teresa PhD Student
