Hi all,
I am solving a series of protein-ligand complex structures in which the
larger ligands typically cause an expansion of the binding site,
changing the receptor into an open conformation, while the smaller
ligands do not change receptor conformation upon binding. In one
structure (1.49 Ang resolution), after several iterations of refinement
with phenix (R-work = 0.1609, R-free = 0.1918), I see both
conformations: all of the closed receptor conformation and the backbone
conformation and 6 out of 7 of the side chains of the open conformation
can be seen in the 2Fo-Fc map (70% occupancy for the closed
conformation, 30% for the open conformation). Modeling in the ligand as
one 100% occupancy pose that fits in the closed receptor conformation
(Fig. 1) gives some negative Fo-Fc density up to sigma = 3.8 and does
not explain the appearance of the open receptor conformation. If I model
the ligand in at 70% occupancy, the negative 2Fo-Fc density is
eliminated and some positive Fo-Fc density appears around the ligand
(Fig.2), suggesting the presence of a second ligand conformation but
there is not enough density to unambiguously place a second ligand
conformation (corresponding to the open receptor conformation). When I
model in a second ligand conformation (Fig. 3), then after refinement
positive Fo-Fc density disappears without any negative Fo-Fc density
appearing but no 2Fo-Fc density appears after refinement to confirm the
correctness of the ligand pose.
Q #1: should I model the ligand in if it eliminates the Fo-Fc positive
density and doesn’t cause negative density? Or should I leave it out
despite the indirect evidence from the altered receptor conformation for
this additional pose? Because this is a series of ligands we know that
the larger receptor conformation implies that a ligand is present in a
pose that opens up the binding site but it's difficult to confirm the
second ligand pose.
Q #2: Is there any way to directly test the correctness of the second
pose that is not dependant on the 2Fo-Fc maps? Is there some kind of
statistical test (such as a local R-factor) to show that the pose is not
in direct disagreement with the data?
A pdf of the figures is available at
http://blur.compbio.ucsf.edu/~merski/figs.pdf
Thanks for your help.
Matthew Merski
UCSF Dept. of Pharmaceutical Chemistry
[email protected]
