Dynamics of an inhibitor at and away from an out-of-active site binding location in human carbonic anhydrase II

The high resolution crystal structure of 2-[(S)-benzylsulfinyl]benzoic acid (3G1)-bound human carbonic anhydrase (HCA) II shows the ligand molecule bound in an out-of-active-site pocket on the surface of HCA II. At this location, it is expected to inhibit the enzyme by arresting the His-64 mediated proton shuttle in its catalytically unimportant outward orientation. However, 3G1 is known to act as a weak inhibitor of HCA II. In this article, we have used computer simulation studies to examine the dynamics of 3G1 molecule for 1 mu s during its long residence in the binding pocket. We have also evaluated the free energy and kinetics of its fast unbinding to the bulk solvent as the 3G1 molecule crosses the bottleneck at the mouth of the binding pocket. Our analysis demonstrates how the dynamical fluctuations around the binding site and unbinding to the bulk solvent render the 3G1 molecule a weak inhibitor of HCA II.