Interior pH-sensing residue of human voltage-gated proton channel Hv1 is histidine 168

Abstract

The molecular mechanisms governing the human voltage-gated proton channel hHv1 remain elusive. Here, we used membrane-enabled hybrid-solvent continuous constant pH molecular dynamics (CpHMD) simulations with pH replica exchange to further evaluate the recently obtained structural models of hHv1 in the closed (hyperpolarized) and open (depolarized) states (Geragotelis, Tobias, et al., Proc. Natl. Acad. Sci. USA 2020) and explore potential pH-sensing residues. The CpHMD titration at a set of symmetric pH conditions revealed three residues that can gain or lose protons upon channel depolarization. Among them, residue H168 at the intracellular end of the S3 helix switches from the deprotonated to the protonated state and its protonation is correlated with the increased tilting of the S3 helix during the transition from the closed to the open state. Thus, the simulation data suggest H168 as an interior pH sensor, in support of a recent finding based on electrophysiological experiments of Hv1 mutants (Cherny, DeCoursey, et al., J. Gen. Physiol. 2018). We propose that protonation of H168 acts as a key that unlocks the closed channel configuration by increasing the flexibility of the S2-S3 linker, which increases the tilt angle of S3 and enhances the mobility of the S4 helix, thus promoting channel opening. Our work represents an important step toward deciphering the pH-dependent gating mechanism of hHv1.