Dear Prem,
If your protein has 2 domains, it is possible that they their relative
orientation is different in your target compared to the search model. Therefore
searching with the two domains separately can improve your Z-score in Phaser
and give you a better map.
However, you did not tell us what your Z-score was after phaser (>= 8.0 after
translation search?) and how your map looks like after phaser, but before
buccaneer. For instance, if you simply refine your initial MR solution in
refmac, do you get density for side chains that are not part of your search
model? Or do you see extra bits of backbone density that are not in the search
model.?What about searching with just one domain, do you see (recognisable)
density for the second?
Finally, an R-value (free or cryst?) of 38% after an initial MR solution is not
unreasonable. You may have to manually rebuild your model in Coot with
subsequent refinement in Refmac to lower your R-values. Of course, rebuilding
in this case requires that you see some extra density you can build into or
some parts of the map where your current model does not fit the density. Have
you tried to do a rigid body refinement of your initial MR solution, where you
allow each domain to move independently?
All the best with this structure.
Klaus
=======================================================================
Dr. Klaus Fütterer
Deputy Head of School
Undergraduate Admissions
Room 717, Biosciences Tower
School of Biosciences P: +44-(0)-121-414 5895
University of Birmingham F: +44-(0)-121-414 5925
Edgbaston E: k.futte...@bham.ac.uk
Birmingham, B15 2TT, UK W: http://tinyurl.com/futterer-lab
=======================================================================
On 2 Dec 2013, at 04:11, Prem Prakash wrote:
> Dear All,
>
> The density obtained after molecular replacement using phaser at 2.5 Angstrom
> and then used buccneer for autobuilding of the model. I am not getting
> reasonable R value (it is 38.5 %) but the figure of merit is 0.629.
>
> As My protein has two domains. So is it possible to fragment the individual
> domain and then again perform the molecular replacement. How it will improve
> my phase more and how R-factor will be reduced ?
>
> Please help and direct me in proceeding in a right way, and if possible
> provide a protocol for doing that. Thank you
>
> With kind regards
>
>
