[phenixbb] Need help in refining my twinned P6322(apparent) dataset
John Pak
john.pak at msg.ucsf.edu
Mon Mar 14 23:55:05 PDT 2011
Hey Phenix community,
Here are the details of my current situation - any assistance in the
matter would be greatly appreciated as I've been struggling mightily for
the past month trying to get this structure refined (to no avail).
Details:
-3.0 angstrom dataset from a membrane protein crystal
-reduced in a P3 Bravis lattice using HKL2000
-Pointless and Xtriage suggest P622 symmetry
-Solved structure via MR - ran Phaser in all P622 based space groups,
found 1 convincing solution in P6322 (LLG ~110) which makes biological
sense (1 monomer in ASU that lies on the screw axis to recapitulate the
biological trimer)
-31% sequence identical search molecule
-no pseudotranslation (via Xtriage)
Xtriage suggests that the P6322 dataset is nearly perfectly twinned
(twin fraction = 0.49). Mindful of the fact that there are no twin
operators for P6322, I'm thinking that the true space group is P321,
P312 or P63 and I've tried a number of things listed below:
1. Refined in P6322 (simulated annealing+individual B). Missing side
chains and incorrectly modelled loops apparent in maps. Start R=0.49,
Rfree=0.5. Refinement stalls at R=0.41, Rfree=0.45 after building.
2. Ran Phaser in P312 (using unrefined model) - no solution.
3. Ran Phaser in P321 (using unrefined model). One nice solution (LLG
~220) - lattice looks very similar to P6322 crystal packing. Twin
refinement in phenix using operator suggested by Xtriage (simulated
annealing + individual B) results in R=0.38, Rfree=0.41 (no rebuilding
performed).
4. Ran Phaser in P63 (using unrefined model). Three solutions (LLGs all
~210) - solution #3 looks very similar to P6322 crystal packing. Twin
refinement in phenix using operator suggested by Xtriage of solution #3
gives R=0.37, Rfree=0.40 (no rebuilding performed). Solutions #1 and #2
do not give packing arrangements that look like the P6322 solution.
Twin refinement of Sol 1 and 2 give higher R factors.
So I'm at a loss as to which space group my crystal truly belongs to,
and where I should be focusing my refinement. The maps all look similar
before and after DM.
Thanks for any assistance or recommendations that you might have!
--
John E. Pak, Ph.D.
Postdoctoral Associate, Stroud Lab
Department of Biochemistry& Biophysics
UCSF MC2240
Genentech Hall Room S414
600-16th St.
San Francisco, CA 94158-2517
Lab #: 415-476-3937
Fax #: 415-476-1902
Cell #: 415-215-0048
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