[ccp4bb]: low res dataset, need help with NCS constraints

[Susan Heffron <sheffron@uci.edu>]

***  For details on how to be removed from this list visit the  ***
***    CCP4 home page http://www.dl.ac.uk/CCP/CCP4/main.html    ***

Hi all!

I could use some advice about refining a model with some fairly low-
resolution data.  Here's my situation:

I have a relatively low resolution dataset (~3.2A) in P212121, which
consists of about 12,000 reflections.  My model is a molecular
replacement solution, with two molecules in the a.u. and a total of
about 6500 atoms.  I have been having trouble getting the refinement
(using CNS) to proceed.  My R-free is still above 30%.  One concern I
have is that the data-to-parameters ratio is less than 1 (12,000
reflections, compared to 19,500 x-y-z positions).  I have been using NCS
restraints, but wonder if constraints would be better, in order to
improve the data-to-parameter ratio.  Am I correct in thinking about it
this way?

If I do use NCS constraints, I have the complication that my protein is
a multi-domain molecule that has flexibility in the orientations of the
domains to each other. So, it would have to be constrained
domain-by-domain.  Which programs can handle this?

One other possibility I've considered is to restrain the structure to a
higher resolution copy with harmonic restraints.  This would require
superimposing a higher-res structure, again domain-by-domain, and then
refitting any loops that are obviously different in the current maps.
(My current structure has a ligand bound, which makes the domain
positions differ from other previous crystal structures even more than
the normal flexibility.) Has anyone had experience with this type of
situation to know whether it would be worthwhile to go through this type
of rebuilding procedure and then restrain it with harmonic restraints
during refinement?

Thanks for any suggestions.

                               Susan Heffron
-- 
------------------------------------
Susan Heffron
Dept. of Physiology & Biophysics
University of California, Irvine
Irvine, CA  92697  U.S.A.

------------------------------------