I've found the following article to be useful in defining suitable refinement strategies at a particular resolution.
1. Mueller, M., Jenni, S. & Ban, N. Strategies for crystallization and structure determination of very large macromolecular assemblies. Curr Opin Struct Biol 17, 572–579 (2007). It can be used as a rough guide line at the start of your refinement (incomplete model, fresh off of molecular replacement) with slow increase of refinement parameters the better your model becomes (and as Rfree permits). F On Mar 2, 2012, at 9:39 AM, Ethan Merritt wrote: > On Friday, 02 March 2012, Regina Kettering wrote: >> Rajesh; >> >> I am not sure that you have a high enough data:refinement parameters ratio >> to refine TLS. >> It just adds more parameters to refine that can lead to over-refinement of >> your model, >> especially at the 3.3 A. > > I'm afraid you've got this completely backwards. > TLS uses very few parameters, and is especially useful at low resolution. > At 3.3A I would recommend trying a TLS model _instead_ of refining > individual B factors. > > NCS restraints also help a lot at low resolution. > > So the drop is believable, but... > > You should first worry about "lots of waters were placed". > It's there that many extra parameters have been added, perhaps leading > to over-fitting. I would not expect 3.3A data to justify placement of > more than a handful of waters at most. > > If you're parameter counting, you might note that 5 water molecules add > more parameters than 1 TLS model. But the TLS model may improve the model > everywhere, whereas the waters will only suppress a few local difference > density peaks. > > cheers, > > Ethan > > >> >> HTH, >> >> Regina >> >> >> >> ________________________________ >> From: Rajesh kumar <ccp4...@hotmail.com> >> To: CCP4BB@JISCMAIL.AC.UK >> Sent: Friday, March 2, 2012 10:54 AM >> Subject: [ccp4bb] sudden drop in R/Rfree >> >> >> >> >> Dear All, >> >> I have a 3.3 A data for a protein whose SG is P6522. Model used was wild >> type structure of same protein at 2.3 A. >> >> After molecular replacement, first three rounds of refinement the R/Rf was >> 26/32.8, 27.1/31.72 % and 7.35/30.88 % respectively. >> In the fourth round I refined with TLS and NCS abd added water and the R/Rf >> dropped to 19.34/26.46. It has almost 7% difference. I also see lot of >> unanswerable density in the map where lot of waters were placed. Model fits >> to the map like a low resolution data with most of side chains don't have >> best density. >> >> I was not expecting such a sudden drop in the R/Rfree and a difference is >> 7.2%. >> I am wondering if I am in right direction. I am not sure if this usual for >> 3.3A data or in general any data if we consider the difference. >> I appreciate your valuable suggestions. >> >> Thanks >> Raj --------------------------------------------- Francis E. Reyes M.Sc. 215 UCB University of Colorado at Boulder
