Dear Ursula,

18% identity is really in the twilight zone for molecular replacement, where it may work but there are certainly no guarantees.

However, there’s a feature in Phaser that is useful for this kind of problem, i.e. the “rotate around” option, where you take advantage of knowing the approximate orientation of a domain.  In the current version of Phaser in either CCP4 or Phenix, this can be done as part of an automated search (instead of doing separate rotation/translation/packing/refinement steps as in earlier versions).  Let’s assume that the model from the first fragment was cut out of a larger model, from which you would now like to place the second fragment.  The rotation to apply to the second one should be similar to what was applied to the first.  In ccp4i, you can go to the “Additional Search Parameters” folder, and choose “Brute” for the “Rotation function target”.  In the “Search” pulldown that appears then, choose “Around an angle”, give the Euler angles that were found in the search for the first domain, and choose something like 20-30 degrees for “Range”, i.e. allow the second domain to differ in orientation by up to 20 or 30 degrees from the orientation for the first domain.  It’s probably better to do translation searches with all orientations generated by this rotation search, so open the “Expert parameters” folder, under “Rotation search peak selection” choose “All peaks”, and turn “Rotation clustering” “Off”.

We’ve had good luck in a number of cases where the hinge angle changes by more than even the best rigid-body refinement would manage.

Good luck!

Randy

-----
Randy J. Read
Department of Haematology, University of Cambridge
Cambridge Institute for Medical Research    Tel: +44 1223 336500
Wellcome Trust/MRC Building                         Fax: +44 1223 336827
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Cambridge CB2 0XY, U.K.                               www-structmed.cimr.cam.ac.uk

On 12 Dec 2014, at 21:38, Ursula Schulze-Gahmen <[log in to unmask]> wrote:

> I am trying molecular replacement with a very poor model. The model consists mainly of 1 long helix and two slightly bent antiparallel helices. After dividing it into 2 fragments, I was able to find a solution for one of the fragments ( at least I think so after looking at maps, packing, refinement etc). But even if I place the first solution as fixed ensemble in phaser, I cannot find a solution for the second fragment ( 18% sequence identity). From the structure of the model and the packing it seems clear where the fragment should go roughly.
> 
> Are there any other programs other than phaser that might be able to solve this problem? I tried already epmr and mr_rosetta without success. 
> I also tried to just superimpose the complete model onto the partial solution. This results in quite nice packing, but doesn't refine. Is there a rigid program refinement program with very large convergence?
> 
> Ursula
> 
> -- 
> Ursula Schulze-Gahmen, Ph.D.
> Project Scientist
> UC Berkeley, QB3
> 360 Stanley Hall #3220
> Berkeley, CA 94720-3220
> (510) 643 9491