[phenixbb] fwd: Re: Need help in refining my twinned P6322(apparent) dataset

Nathaniel Echols nechols at lbl.gov
Tue Mar 15 09:03:32 PDT 2011

---------- Forwarded message ----------
From: "Van Den Berg, Bert" <Lambertus.VandenBerg at umassmed.edu>
To: "PHENIX user mailing list" <phenixbb at phenix-online.org>
Date: Tue, 15 Mar 2011 08:01:42 -0400
Hi John,

How do the maps look for cases 3 and 4? Are they great or are there
obvious things to correct? Does your model have the correct sequence
or is it that of the search model? I’m asking because your R’s seem
reasonable to me after a first round of refinement and using a 30%
identical search model for MR. So maybe there is no problem?


On 3/15/11 2:55 AM, "John Pak" <john.pak at msg.ucsf.edu> wrote:

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).


-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

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

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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