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You've probably done it already, but solvent flattening/flipping/massaging at 80% solvent should provide a cheap thrill with regard to phase improvement. ===================================== Phoebe A. Rice Dept. of Biochemistry & Molecular Biology The University of Chicago phone 773 834 1723 http://bmb.bsd.uchicago.edu/Faculty_and_Research/01_Faculty/01_Faculty_Alphabetically.php?faculty_id=123 http://www.rsc.org/shop/books/2008/9780854042722.asp ---- Original message ---- >Date: Fri, 28 Jan 2011 14:29:07 -0600 >From: CCP4 bulletin board <[log in to unmask]> (on behalf of Todd Geders <[log in to unmask]>) >Subject: [ccp4bb] Noisy difference maps with high solvent content? >To: [log in to unmask] > > Greetings CCP4bb, > > Short version: > > Very noisy difference maps from a crystal with > extremely high solvent content, seeking advice on > how best to handle such high solvent content to > eliminate noise in difference maps. > http://strayxray.com/images/coot.jpg > > Long version: > > I'm having trouble with a 3.0Å dataset from a > crystal with 80% solvent content. The space group > is P4132 and I'm quite confident the high solvent > content is real (there is a species-specific set of > helices extending into the solvent channels that > appears to prevent tighter packing). > > I was able to get a MR solution using a structurally > related enzyme, but the difference maps are terribly > noisy (see link). There are lots of negative > density in empty spaces between well-defined 2Fo-Fc > electron density. > > http://strayxray.com/images/coot.jpg > > The 2Fo-Fc density actually looks fairly good. The > initial MR maps had clear density correlating to the > sequence differences between the MR model and the > crystallized protein. After fixing the model as > best I could, the refinement statistics are R/Rfree > of 27.5/30.3 with a data/parameters ratio of 1.7. > > The mosaicity ranges from 0.15-0.3, data were > collected with 0.5° oscillations and 180° of data > were collected. > > http://strayxray.com/images/diffraction.jpg > > Since the crystals appeared to suffer from radiation > decay (based on scaling statistics), I only use the > first 40° of data (which still gives around 8-fold > redundancy). Using more minimal wedges of data or > more data does not noticeably make better or noisier > maps. > > Any advice on improving the maps? Could the noisy > maps be due to the extraordinarily high solvent > content? > > I'd appreciate any advice or comments. > > ~Todd Geders