[phenixbb] Molecular replacement at low resolution

Gino Cingolani Gino.Cingolani at jefferson.edu
Sat May 2 09:18:32 PDT 2015

Here what you could try:

First, make sure your data are not twinned.

Second (assuming your data are not twinned), use Self Rotation function (I like GLRF) to determine the exact number of copies in the asu. A search for k=45 should 'show' the existence and orientation of a 8-fold ncs axis. If you don't see anything at k=45, then you don't have 8 copies in the asu. Try different options (e.g. k=60 for 6-fold, k=90 for 4-fold, etc), ultimately k=180 should give you an equal number of 2-fold axes as the number of particles in the asu.

Third, try MR with whatever 'geometric' model you have (better smaller then larger).

Don't be bothered by low LLG values.If you have a solution that 'makes sense' (packing-wise)

compute self rotation functions with Icalc and compare them to those calculated with Iobs.

The two should look alike, if the solution is correct.

Fourth, assuming the above have worked, you need to go from a 'geometric' model to a

'physical entity' to calculate phases. Mask you protomer and bootstrap real phases

by ncs-averaging density modification (e.g. resolve, dm, etc).

Fifth, calculate an averaged map, build/improve your initial model and used Pavel's new real_space_refinemnet module to refine it against the averaged density.

Real space refinement has larger radius of convergence that classical rigid/positional refinement

and works much better at medium resolution (~4A).

Good luck!



Gino Cingolani, Ph.D.
Thomas Jefferson University
Dept. of Biochemistry & Molecular Biology
233 South 10th Street - Room 826
Philadelphia PA 19107
Tel: (215) 503 4573
Website:  http://www.cingolanilab.org
"Nati non foste per viver come bruti, ma per seguir virtute e canoscenza"
("You were not born to live like brutes, but to follow virtue and knowledge")
Dante, The Divine Comedy (Inferno,  XXVI, vv. 119-120)

From: phenixbb-bounces at phenix-online.org <phenixbb-bounces at phenix-online.org> on behalf of Roger Rowlett <rrowlett at colgate.edu>
Sent: Saturday, May 02, 2015 11:50 AM
To: mohamed noor
Cc: PHENIX user mailing list
Subject: Re: [phenixbb] Molecular replacement at low resolution

Before giving up, search in all 8 possible P422 space groups, individually if necessary. You may have to relax or eliminate packing restraints to get solutions or partial solutions to inspect. Looking at packing of these solutions may give you some clues about the true number of molecules in the ASU and eliminate dome space groups. Matthews calculations are not terribly reliable for N>2. Treat your 8 as a very rough estimate.

If Phaser doesn't work, try EPMR. Ultimately, you may not have sufficient data, but we just solved a difficult data set in P222 point group at 3.4 A with N=6, (Matthews predicted N=8). Looking at packing in Coot suggested a packing problem where a tetramer straddled one of the 2 fold axes. Searching for 3 dimers worked nicely.

Roger Rowlett
Gordon & Dorothy Kline Professor
Department of Chemistry
Colgate University

On May 2, 2015 11:01 AM, "mohamed noor" <mohamed.noor34 at gmail.com<mailto:mohamed.noor34 at gmail.com>> wrote:
Dear developers

I have a dataset to about 4 A in point group P 4 2 2 (Pointless suggests P 41 21 2). The protein is about 11 kDa and has an identity of about 50 % to my search model. Phaser calculated the eLLG to be about 5, and maybe not surprisingly, it failed even after using Sculptor to prepare the model.

It also says that solutions with Z-scores more than 8 were rejected for failing the packing test. Are there any suggestions on getting a solution?

Solvent content calculation from Xtriage suggests about 8 copies in the asu. Would this be of any help in MR?


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