Hi Wulf, For your waters that move slightly out of the centre of density I think SHELX FAQ #27 would apply:
Q27: Some Waters are moving away from the centre of density toward the corner of the density. What is going on ? George: We have sometimes seen atoms move to the edge of density in a high resolution refinement. We attribute this to (a) Fourier series termination errors because of missing reflections (including Rfree reflections), and (b) the weights for refinement are of necessity different to those used to make a Fourier map (which theoretically requires unit weights). In either case these refined positions are the 'correct' ones, the map is misleading. However anti-bumping (BUMP) and other geometric restraints can also move atoms out of density, in this case you need to check whether you need to include partial occupancies. BUMP is not applied if the occupancy sum is less than 1.1. I have in the past been comforted by this statement by George... You can also try the SHELXWAT module. Cheers, Martin . Martin Hallberg, PhD Department of Cell and Molecular Biology Karolinska Institutet Stockholm Sweden http://tinyurl.com/yzn9y5j On Nov 12, 2009, at 8:53 AM, Wulf Blankenfeldt wrote: > Dear all, > > I am blessed/tormented with a 0.8 A data set for a small protein. I first > refined it using COOT/REFMAC5 until I deemed it done, which meant manually > adding quite a few water molecules that were weak and a little more distant > to the protein. R is 10.2%, R-free is 10.8%. > > I realize that referees will probably ask for a SHELXL refinement, so I spent > the last few days learning about FVAR and SUMP instructions and the like and > finally got it running in a way I thought should be correct. Using my final > REFMAC5 model, I get R=10.8% and R-free=11.1% - I can certainly live with > this, but: > > When I now look at the electron density, a large number of my tenderly > curated water molecules have lost their 2FO-FC electron density (50 out of > 200; I usually use a 1 sigma cutoff), or they have shifted to lie beside a > density blob. It may have to do with the fact that I assigned 0.5 occupancy > to some of them and now they have been BUMPed away, others may simply be too > weak to stay in place. > > I wonder what to do now. Do I have to rebuild the water structure again and > refine everything in SHELXL exclusively? Or are there any flags I could use > to keep the old waters intact (risking higher R-factors)? > > Any help is highly appreciated! > > Thank you in advance, > > > Wulf
