Thank you both, Mischa and Harry, I'll try overall B refinement. But I'm starting to think that my Refmac is somewhat crappy, I tried this before, and then all the B-factors were exactly the same. Now, I tried it again and they are not... Something's still wrong here.
(But I'm still curious about the B-factor refinement when there is no "REFI BREF ISOT" in the com-file...) Eva 2007/4/18, Mischa Machius <[EMAIL PROTECTED]>:
Like Harry said, it is not justified to do individual B factor refinement at that resolution. Well, you can do it, but you'll end up with funny results, such as what are observing right now. Still, from a pragmatic point of view, individual B factor refinement in cases like these can have a positive effect on the electron density. However, keep in mind that the resulting B factors may physically not be very meaningful. In the end, you'll have to switch to grouped B factor refinement, or you risk nasty comments from an attentive mentor or reviewer (and rightly so). Hope that helps. Best - MM On Apr 18, 2007, at 7:20 AM, Eva Kirchner wrote: Hi, I have a little problem with B-factor refinement. I'm using the CCP4i interface, Refmac 5.2.0019, a resolution of 30-3.2 A (I tried 8-3.2 A as well, it doesn't make a big difference for this problem), and a current Rfree of 30.4%. Refmac refines the B-factors so that they are nearly the same for main chain and side chain, and I don't like that (or could it make sense in any way?). Moreover, my structure is a protein complex, and Refmac is mainly doing this for one component of the complex. If I take the B-factors from the original uncomplexed protein (around 18, 1.75 A) and add 44 to them with moleman to get them in the range they are in the complex, Refmac "flattens" them remarkably in only 5 cycles of restricted refinement. Does anyone have an explanation for this? I am pretty sure that the complex components are in the right place, I see beautiful density and everything I should see at this resolution. Here is what I tried further: * I de-selected "Refine isotropic temperature factors" in the Refmac interface. There was no REFI BREF ISOT any more in the com file. But there was also no difference in the B-factors compared to when there _was_ REFI BREF ISOT in the com file... So does Refmac just _ignore_ my wish not to refine B-factors? (The REFI keywords were as follows: type REST - resi MLKF - meth CGMAT - is there any B-factor-thing hidden in this?) * I played around with the geometric parameters. If I select the B-factor values there (the keywords are TEMP|BFAC <wbskal><sigb1><sigb2><sigb3><sigb4>), it does not make _any_ difference, what values I fill in there, the resulting B-factors are always the same (but different from when I don't use the TEMP keyword, and even "flatter"). Default for WBSCAL is 1.0, I tried 10, 1.0, 0.1, 0.01, and the equivalent numbers for the sigbs. Thanks for any thoughts on this, Eva -------------------------------------------------------------------------------- Mischa Machius, PhD Associate Professor UT Southwestern Medical Center at Dallas 5323 Harry Hines Blvd.; ND10.214A Dallas, TX 75390-8816; U.S.A. Tel: +1 214 645 6381 Fax: +1 214 645 6353
