Properties of the Voltage-Gated Proton Channel Gating Currents

Abstract

The voltage-gated proton channel (Hv1) is a dimer of subunits containing four transmembrane domains (S1-S4) which play the double role of harboring the voltage-sensing charges and forming the proton conduction pathway. Some biophysical properties of Hv1, as the coupling between voltage-sensing and permeation pathway opening, the pH dependence of gating, the cooperativity between each subunit in the dimer, and the steps of charge movement during activation and deactivation can be better understood by measuring and characterizing the Hv1 gating currents. However, to measure gating currents, ionic currents needs to be eliminated or greatly reduced, a feat that is particularly challenging for Hv1 for which there is no effective blocker and proton concentration cannot be reduced more than 10 fold. Here, we expressed the low-conducting N264R Hv1 mutant from Ciona intestinalis in Xenopus laevis oocytes and succeeded in recording gating currents membrane patches with both, the monomeric and dimeric channels. In agreement with previous fluorescence measurements (Qiu et al., Neuron 77:288, 2013), the analysis of the gating currents kinetics and charge movement revealed that the ON component has a double exponential decay. Additionally, changes in pH modulate Hv1 gating currents kinetics and Q-V curves. This suggests that the pH sensors in Hv1 are located in the voltage-sensing domain. Finally, in our experimental conditions we observed charge immobilization after activation, which could be an intrinsic characteristic of Hv1 voltage-sensing domain. Supported by CONICYT-PFCHA/Doctorado Nacional/2017-21170395 to E.C., Fondecyt grants ACT 1104, Fondecyt 1160261 to C.G. and 1150273 to R.L.