I got good data to 2.4A on a protein-ligand complex. I want to solve the
structure by MR using a model with 45% seq. ID and 55% similarity.
Initially the data appeared to be P622 (pointless, selfrotation function
etc). Unit cell: 145 145 65, MW protein is 28kDa
I prepared the MR model with chainsaw, choping of non-conserved residues
at the C gamma and reset the B-factors. Phaser found a solution, but
with negativ LLG.
I than processed the data in P3, P321, P312 and P6 and run Phaser
searching all possible alternative spacegroups. The best solution is P3
(4mol/asu) followed by P321 (2mol/asu),both with a LLG of above 400. But
when trying to refine the models in Refmac the R/Rfree stays at ~48%.
Looking at the truncate output and phenix xtriage, twinning is suggested
with the merohedral twin law -h, -k, l and a twin fraction of 40.3%
Using twin refine option in Refmac (5.5.0070), the R/Rfree for the P321
solution drops to around 33%, but the difference between R/Rfree is only
1%. For the solution found in P3 the R/Rfree stays at around 47%.
So I assumed that P321 is the better solution. Is the difference in
R/Rfree only 1%, because the free and work reflections are related
through the twin law?
I used phenix to assign the free reflections putting in the twin
operators. Doing simulated annealing in phenix, I get a rather large
difference in R/Rfree of ~7%. Well, I guess I need to do some tweaking
of the parameters in phenix (running the latest phenix and cci_apps).
When I than use these free reflections assigned by phenix in Refmac I
still get only 1.5% difference between R and Rfree?
So is it doable in Refmac, or is my best bet phenix? Any advice what is
the best way to proceed is much appreciated!
Dr. Sabine Schneider
Department of Chemistry and Pharmacy
Butenandtstrasse 5-13, Building F
Phone: +49 (0)89 2180 77846
Fax: +49 (0)89 2180 77756