Hi Roger,
personally, I would take the four dimers from Phaser, strip off all side
chains (make a poly-ala/gly), do a ML refinement with tight NCS
restraints. The resulting map could then be on-the-fly
real-space-averaged with Coot. At your resolution, I would expect to see
the real register of at least some side chains either in the 2mfodfc-map
or in one of the real-space-averaged 2mfodfc-maps.
Good luck,
Dirk.
Am 01.09.10 17:12, schrieb Roger Rowlett:
I am trying to find a MR solution for a large unit cell (R3:H,
158x158x196) with a relatively poor, but I think workable search model
(30% identity, 50% similarity). The data set is decent to 2.4 A. I
might be able to get better if necessary. I submitted the sequence of
the target to the Phyre server, and it returned a PDB file derived
from what I had already identified as the most likely successful
search model. Phaser finds a reasonable solution with four dimers
(Z=9-15 depending on the data set) for which the unit cell packing
looks good. Phaser even assembles what looks like "correct" biological
tetramers in the ASU and across the symmetry interfaces. The main
chain appears to be mostly traceable, but the side chains are not all
well-resolved, and I suspect from the better-defined regions, that the
chain is misregistered by a residue or two throughout most of the
structure.
Assuming that an MR solution is possible, what is a good approach from
here? FWIW, Phaser does not correctly place a poly-Ala/Gly model,
although it may place three chains similarly to the MR solution I have
with the Phyre model. My first thought is to:
1. Chainsaw the currrent solution, and attempt to identify and build
in the correct register of the side chains. after refinement.
2. Do a low resolution refinement of the poly-Ala/Gly model to better
thread the main chain?
3. Try EPMR with the Phyre model or the poly-Ala/Gly model
4. Give up and get real phases (but I'm so close now!?)
Thanks in advance.
--
Roger S. Rowlett
Professor
Department of Chemistry
Colgate University
13 Oak Drive
Hamilton, NY 13346
tel: (315)-228-7245
ofc: (315)-228-7395
fax: (315)-228-7935
email: rrowl...@mail.colgate.edu <mailto:rrowl...@mail.colgate.edu>
--
*******************************************************
Dirk Kostrewa
Gene Center Munich, A5.07
Department of Biochemistry
Ludwig-Maximilians-Universität München
Feodor-Lynen-Str. 25
D-81377 Munich
Germany
Phone: +49-89-2180-76845
Fax: +49-89-2180-76999
E-mail: kostr...@genzentrum.lmu.de
WWW: www.genzentrum.lmu.de
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