Allosteric Inhibitors Have Distinct Effects, but Also Common Modes of Action, in the HCV Polymerase

Abstract

The RNA-dependent RNA polymerase from the Hepatitis C Virus (gene product NS5B) is a validated drug target because of its critical role in genome replication. There are at least four distinct allosteric sites on the polymerase to which several small molecule inhibitors bind. In addition, numerous crystal structures have been solved with different allosteric inhibitors bound to the polymerase. However, the molecular mechanisms by which these small molecules inhibit the enzyme have not been fully elucidated. There is evidence that allosteric inhibitors alter the intrinsic motions and distribution of conformations sampled by the enzyme. In this study we use molecular dynamics simulations to understand the structural and dynamic changes that result when inhibitors are bound at three different allosteric binding sites on the enzyme. We observe that ligand binding at each site alters the structure and dynamics of NS5B in a distinct manner. Nonetheless, our studies also highlight commonalities in the mechanisms of action of the different inhibitors. Each inhibitor alters the conformational states sampled by the enzyme, either by rigidifying the enzyme and preventing transitions between functional conformational states or by destabilizing the enzyme and preventing functionally relevant conformations from being adequately sampled. By illuminating the molecular mechanisms of allosteric inhibition, these studies delineate the intrinsic functional properties of the enzyme and pave the way for designing novel and more effective polymerase inhibitors. This information may also be important to understand how allosteric regulation occurs in related viral polymerases and other enzymes.