Structural Modeling of Local Anesthetic Binding to the Pore-Domain of Human Nav1.5 in Open and Closed States using Rosetta

Abstract

Mutations in voltage-gated sodium (Nav) channel isoforms are correlated with a wide range of cardiovascular and neurological diseases in humans, and are therefore are important targets for the rational design of novel drugs. The cardiac isoform of the Nav channel, Nav1.5, presents a unique target for the development of antiarrhythmic drugs. In this work, we identify key structural motifs for local anesthetic binding in the pore-domain of the open and closed states of the human Nav1.5 channel. The Rosetta structural modeling method was used to construct models of human Nav1.5 isoform based on the 3D crystal structures of bacterial Nav channels: NavRh (closed state) and NavMs (open state). The resulting lowest free-energy models were selected for local anesthetic docking simulations. Rosetta loop modeling and global relaxation of 10,000 models yielded a convergent motif in the selectivity filter region of a stabilizing hydrogen bond network between Tryptophan and Threonine pairs. A membrane-facing fenestration near the S6 helix of domain IV and the S5 helix of domain III was also structurally conserved, and is a proposed site of neutral drug entry. Docking simulations of the channel blocker, lidocaine, reveal key protein-ligand binding configurations within the pore. Our preliminary models of the human Nav1.5 channel in the open and closed states reveal highly conserved structural motifs important for both stabilization of the pore domain, as well as for drug entry and binding. Future work will use structural models of drug interaction with human Nav1.5 as a dynamic testing platform for the calculation of the kinetics of drug binding and unbinding.