Rationally Designed Proton Channel Inhibitors Reveal a Druggable Pocket in a Voltage-sensing Domain

Abstract

Voltage-gated sodium, potassium, and calcium channels play critical roles in excitable tissues, from the generation and propagation of action potentials to synaptic transmission and muscle contraction. As a result, they are pharmacological targets for the treatment of a variety of diseases. They consist of four voltage-sensing domains (VSDs) surrounding a central pore domain. While many types of organic molecules bind pore domains, the number of molecules known to bind VSDs is limited. The proton channel Hv1 is made of two VSDs and lacks a pore domain, providing a simplified model for studying how small ligands interact with VSDs. We previously identified a binding site for arginine-mimic compounds in the center of the Hv1 VSD, which is accessible only when the proton-conduction pathway is open. Here we show that a new generation of rationally designed arginine mimics, named HIFs, are able to interact with an additional binding pocket within the VSD intracellular vestibule. HIFs can reach the binding pocket even when the proton-conduction pathway is closed. Once inside the pocket, they become trapped and can only be released over extended periods of time (several minutes). We used electrophysiological measurements, combined with kinetic modeling, molecular docking, and atomistic simulations to determine the location and composition of the pocket. Our findings suggest that similar binding sites could be found in the VSDs of other channels and exploited for drug development.