[phenixbb] occupancy refinement

Pavel Afonine PAfonine at lbl.gov
Tue Nov 24 20:37:16 PST 2009

Hi Maia,

the atom parameters, such as occupancy, B-factor and even position are 
interdependent in some sense. That is, if you have somewhat incorrect 
occupancy, that B-factor refinement may compensate for it; if you 
misplaced an atom the refinement of its occupancy or/and B-factor will 
compensate for this. Note in all the above cases the 2mFo-DFc and 
mFo-DFc maps will appear almost identical, as well as R-factors.

So, I think your goal of finding a "true" occupancy is hardly achievable.

Although, I think you can approach it by doing very many refinements 
(say, several hundreds) (where you refine occupancies, B-factors and 
coordinates) each refinement starting with different occupancy and 
B-factor values, and make sure that each refinement converges. Then 
select a subset of refined structures with similar and low R-factors 
(discard those cases where refinement got stuck for whatever reason and 
R-factors are higher) (and probably similar looking 2mFo-DFc and mFo-DFc 
maps in the region of interest). Then see where the refined occupancies 
and B-factors are clustering, and the averaged values will probably give 
you an approximate values for occupancy and B. I did not try this myself 
but always wanted to.

If you have a structure consisting of 9 carbons and one gold atom, then 
I would expect that the  "second digit" in gold's occupancy would 
matter. However, if we speak about dozen of ligand atoms (which are 
probably a combination of C,N,O) out of a few thousands of atoms of the 
whole structure, then I would not expect the "second digit" to be 
visibly important.


On 11/24/09 8:08 PM, chern wrote:
> Thank you Kendall and Pavel for your responces.
> I really want to determine the occupancy of my ligand. I saw one 
> suggestion to try different refinements with different occupancies and 
> compare the B-factors.
> The occupancy with a B-factor that is at the level with the average 
> protein B-factors, is a "true" occupancy. 
>     I also noticed the dependence of the ligand occupancy on the 
> initial occupancy. I saw the difference of 10 to 15%, that is why I am 
> wondering if the second digit after the decimal point makes any sence.
> Maia
>     ----- Original Message -----
>     *From:* Kendall Nettles <mailto:knettles at scripps.edu>
>     *To:* PHENIX user mailing list <mailto:phenixbb at phenix-online.org>
>     *Sent:* Tuesday, November 24, 2009 8:22 PM
>     *Subject:* Re: [phenixbb] occupancy refinement
>     Hi Maia,
>     I think the criteria for occupancy refinement of ligands is
>     similar to a decision to add an alt conformation for an amino
>     acid. I  don't refine occupancy of a ligand unless the difference
>     map indicates that we have to. Sometimes part of the igand may be
>     conformationally mobile and show poor density, but I personally
>     don't think this justifies occupancy refinement without evidence
>     from the difference map. I agree with Pavel that you shouldn't
>     expect much change in overall statistics, unless the ligand has
>     very low occupancy., or you have a very small protein.  We
>     typically see 0.5-1% difference in R factors from refining with
>     ligand versus without for nuclear receptor igand binding domains
>     of about 250 amino acids, and we see very small differences from
>     occupancy refinement of the ligands.
>     Regarding the error, I have noticed differences of 10% percent
>     occupancy depending on what you set the starting occupancy before
>     refinement. That is, if the starting occupancy starts at 1, you
>     might end up with 50%, but if you start it at 0.01, you might get
>     40%. I don't have the expertise to explain why this is, but I also
>     don't think it is necessarily important. I think it is more
>     important to convince yourself that the ligand binds how you think
>     it does. With steroid receptors, the ligand is usually planer, and
>     tethered by hydrogen bonds on two ends. That leaves us with with
>     four possible poses, so if in doubt, we will dock in the ligand in
>     all of the four orientations and refine. So far, we have had only
>     one of several dozen structures where the ligand orientation was
>     not obvious after this procedure. I worry about a letter to the
>     editor suggesting that the  electron density for the ligand
>     doesn't support the conclusions of the paper, not whether the
>     occupancy is 40% versus 50%.
>     You might also want to consider looking at several maps, such as
>     the simple or simulated annealing composite omit maps. These can
>     be noisy, so also try the kicked maps (
>     http://www.phenix-online.org/pipermail/phenixbb/2009-September/002573.html),
>     <http://www.phenix-online.org/pipermail/phenixbb/2009-September/002573.html%29,>
>     which I have become a big fan of.
>     Regards,
>     Kendall Nettles
>     On 11/24/09 3:07 PM, "chern at ualberta.ca" <chern at ualberta.ca> wrote:
>         Hi,
>         I am wondering what is the criteria for occupancy refinement of
>         ligands. I noticed that R factors change very little, but the
>         ligand
>         B-factors change significantly . On the other hand, the
>         occupancy is
>         refined to the second digit after the decimal point. How can I
>         find
>         out the error for the refined occupancy of ligands?
>         Maia
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