Histidine(168) is crucial for ΔpH-dependent gating of the human voltage-gated proton channel, hH(V)1

We recently identified a voltage-gated proton channel gene in the snail Helisoma trivolvis, HtH(V)1, and determined its electrophysiological properties. Consistent with early studies of proton currents in snail neurons, HtH(V)1 opens rapidly, but it unexpectedly exhibits uniquely defective sensitivity to intracellular pH (pH(i)). The H(+) conductance (g(H))-V relationship in the voltage-gated proton channel (H(V)1) from other species shifts 40 mV when either pH(i) or pH(o) (extracellular pH) is changed by 1 unit. This property, called ΔpH-dependent gating, is crucial to the functions of H(V)1 in many species and in numerous human tissues. The HtH(V)1 channel exhibits normal pH(o) dependence but anomalously weak pH(i) dependence. In this study, we show that a single point mutation in human hH(V)1—changing His(168) to Gln(168), the corresponding residue in HtH(V)1—compromises the pH(i) dependence of gating in the human channel so that it recapitulates the HtH(V)1 response. This location was previously identified as a contributor to the rapid gating kinetics of H(V)1 in Strongylocentrotus purpuratus. His(168) mutation in human H(V)1 accelerates activation but accounts for only a fraction of the species difference. H168Q, H168S, or H168T mutants exhibit normal pH(o) dependence, but changing pH(i) shifts the g(H)-V relationship on average by <20 mV/unit. Thus, His(168) is critical to pH(i) sensing in hH(V)1. His(168), located at the inner end of the pore on the S3 transmembrane helix, is the first residue identified in H(V)1 that significantly impairs pH sensing when mutated. Because pH(o) dependence remains intact, the selective erosion of pH(i) dependence supports the idea that there are distinct internal and external pH sensors. Although His(168) may itself be a pH(i) sensor, the converse mutation, Q229H, does not normalize the pH(i) sensitivity of the HtH(V)1 channel. We hypothesize that the imidazole group of His(168) interacts with nearby Phe(165) or other parts of hH(V)1 to transduce pH(i) into shifts of voltage-dependent gating.