[phenixbb] Estimation of ligand binding affinity by occupancy refinement

[Zoran Stefanic]

Dear all,

I have a question about sensitivity of occupancy refinement in phenix. I 
have a series of datasets done at our home instrument of relatively 
modest resolutions from 2.8 to 3 A. These are performed on crystals 
soaked in progressively higher ligand concentrations. We are trying to 
see to which of the several active sites the ligand binds first so we 
went from low to high ligand concentrations. We wanted to estimate the 
amount of ligand bound by refining the occupancies of the ligand. We 
expect one active site to bind more strongly than the other two active 
sites. This was indeed what we observed but when we refine the 
occupancies of the ligands in weaker binding sites starting at 0.2 
occupancies they rise to 0.5-0.6 although some atoms are totally out of 
density and some are in. B factors are reasonable, i.e. don't go wild.

My question would be: how reliable is this estimation of ligand binding 
by occupancy refinement? Why doesn't the occupancy of ligands stay low 
if afterwards the density fit of the ligand looks rather unconvincing?

Thank you in advance
Zoran

-- 
Dr. Zoran Stefanic

Division of Physical Chemistry
Rudjer Boskovic Institute
Bijenicka 54
P.O. Box 180
Zagreb
Croatia

Phone: 00 385 1 457 1203
Fax: 00 385 1 468 0245