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 hH v 1 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 hH v 1 in hyperpolarized (closed channel) and depolarized (open channel) 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 hyper-to the depolarized state. Thus, the simulation data suggest H168 as an interior pH sensor, in support of a recent finding based on electrophysiological experiments of H v 1 mutants (Cherny, DeCoursey et al., J. Gen. Physiol. 2018). Our work represents an important step towards deciphering the pH-dependent gating mechanism of hH v 1. SIGNIFICANCE The human voltage-gated proton channel hH v 1 is comprised of a proton-selective voltage sensing domain and responsible for cellular pH homeostasis. Despite intense experimental and theoretical investigations, its pH-dependent gating mechanism is not understood. Our simulation data offer strong evidence supporting the role of H168 as a pH i sensor (Cherny, DeCoursey et al., J. Gen. Physiol. 2018). Deciphering the interior pH sensor moves us a step closer to elucidating the structure-function relationship of hH v 1.