Gating Currents in the Hv1 Proton Channel

Abstract

The Hv1 proton channel shares striking structural homology with fourth transmembrane helical segment-type voltage-sensor (VS) domains but manifests distinctive functional properties, including a proton-selective "aqueous" conductance and allosteric control of voltage-dependent gating by changes in the transmembrane pH gradient. The mechanisms responsible for Hv1's functional properties remain poorly understood, in part because methods for measuring gating currents that directly report VS activation have not yet been described. Here, we describe an approach that allows robust and reproducible measurement of gating-associated charge movements in Hv1. Gating currents reveal that VS activation and proton-selective aqueous conductance opening are thermodynamically distinct steps in the Hv1 activation pathway and show that pH changes directly alter VS activation. The availability of an assay for gating currents in Hv1 may aid future efforts to elucidate the molecular mechanisms of gating cooperativity, pH-dependent modulation, and H selectivity in a model VS domain protein.

On the control of the proton current in the voltage-gated proton channel Hv1

“…Using our more microscopic direct calculation of the gating charge (24) gives a gating charge of 1.6 (e 0 ). Although we underestimate the observed Q, we note that the analysis of the gating charge is not unique; and depends on the fitting and the pH (37). Furthermore, the model may reflect the effect of the membrane surface charges that should have destabilized the ionized acids on the inside and neutralized some of them.…”

Gating charge displacement in a monomeric voltage-gated proton (H v 1) channel

“…Surprisingly, the gating current data do not show a Cole-Moore effect like the one reported here. This is an unexpected result, considering that the data also indicate that the gating current kinetics is defined by multiple states (41). However, the lack of a well-defined Cole-Moore effect may be due to the fast activation kinetics conferred by the W207A mutation (42).…”

“…Note. While this paper was under revision, detection of gating currents in the dimeric human proton channel mutant W207A-N214R (W257A-N264R in Ciona-H v 1) was reported (41). The method used by the authors to measure gating currents was stepping to a depolarizing voltage from different holding potentials, which is essentially a "Cole-Moore" protocol.…”

Voltage vs. Ligand I: Structural basis of the intrinsic flexibility of S3 segment and its significance in ion channel activation

“…Whereas CNG channels are almost completely dependent on cyclic nucleotides to be activated [67], TRPV channels are frequently considered as polymodal receptors responding to a plethora of stimuli, including a diverse spectrum of natural compounds with agonistic activity [9]. However, channels with high rigidity or intermediate flexible S3 profiles (K V 7, K V 10-12, Hv1) are typically gated only by voltage and just modulated by specific ligands such as divalent cations, protons and phosphoinositides [68][69][70]. On the other hand, channels exhibiting a highly flexible VSD, such as CNG or the Ca 2+ -activated BK Ca channels (Slo or MaxiK channels), are sensitive to the membrane potential in a lesser extent than those channels with a more rigid profile, belonging to the K V or EAG families, according to the reported elementary charges moving through the CTC during activation (Table 1).…”