[phenixbb] high average b-factor vs. Wilson B

Gino Cingolani Gino.Cingolani at jefferson.edu
Wed Aug 4 05:36:46 PDT 2010

Dear Geoffrey,

I had similar issues on a lower resolution structure, which Phenix kept refining to a B-factor >120-150A2, despite my average Wilson B-factor was (only) 85A2. 

I am aware of all approximation made in defining the Wilson B-factor and how one should not take the Wilson B-factor too literally (etc.. etc...). But, as a matter of fact, Phenix kept over-refining my B-factor to crazy values. And I have seen in 
two cases, both with VERY large structures with >15,000 residues/asym unit and ncs ranging between 4- to 24-fold. 
So I admit these structures may not be your everyday refinement  benchmark. At any rate, after several months of troubleshooting here what I can suggest:


1. Was your diffraction anisotropic? If so, try the 
Diffraction Anisotropy Server. The server applies a negative isotropic B-factor correction to your data. In my case, corrected structure factor amplitudes resulted in a 2.5% drop in the Rfree and the average B-factor of my structure (which phenix routinely refined to >120-150A2 after only 3 macrocycles) went down to ~90A2, quite close to my Wilson B-factor.
More importantly, when I use 'corrected' data, I can refine 
even with 10 macrocycles and my structure B-factor stays constant.

How many macrocycles of refinement are you using?
In my case Phenix kept bumping up the average B-factor of my structure by ~10A2 macrocycle. A 10 macrocycles refinement would bring the averaged B-factor to ~200A2. So don't over-refine.
Stick to 2-3 cycles (provided your geometry is good, of course).

Do you have NCS and are you using TLS?
If so, try to first use NCS to refine atom positions, 
then do TLS refinement without imposing NCS. 
If you think about, it doesn't make too much sense to use ncs-restraints when splitting a structure in TLS groups.

Are you playing with weights (say ... wxc_scale=0.5 and wxu_scale=1) and do you have NCS? If so, tightening the geometry too much in the presence of NCS can bump-up our B-factor. 
Inspect your Phenix output file to look at the rms deviations among your ncs-copies in the asymmetric unit.

Good luck!


Gino Cingolani, Ph.D.
Associate Professor
Thomas Jefferson University
Dept. of Biochemistry & Molecular Biology
233 South 10th Street - Room 826
Philadelphia PA 19107
Office (215) 503 4573
Lab    (215) 503 4595
Fax    (215) 923 2117
E-mail:   gino.cingolani at jefferson.edu
"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) 

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