Pavel, my question is this. If occupancy depends on
B-factor, how come the occupancy refinement gives you such an accurate number
for an occupancy (let's say 0.44).
Maia
----- Original Message -----
Sent: Tuesday, November 24, 2009 9:08
PM
Subject: Re: [phenixbb] occupancy
refinement
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 -----
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),
which I have become a big fan of.
Regards,
Kendall
Nettles
On 11/24/09 3:07 PM, "chern@ualberta.ca"
<chern@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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