Structural dynamics determine voltage and pH gating in human voltage-gated proton channel
- University of Missouri-Kansas City, United States
Abstract
Voltage-gated ion channels are key players of electrical signaling in cells. As a unique subfamily, voltage-gated proton (Hv) channels are standalone voltage sensors without separate ion conductive pores. Hv channels are gated by both voltage and transmembrane proton gradient (i.e ∆pH), serving as acid extruders in most cells. Amongst their many functions, Hv channels are known for regulating the intracellular pH of human spermatozoa and compensating for the charge and pH imbalances caused by NADPH oxidases in phagocytes. Like the canonical voltage sensors, Hv channels are a bundle of 4 helices (named S1 through S4), with the S4 segment carrying 3 positively charged Arg residues. Extensive structural and electrophysiological studies on voltage-gated ion channels, in general, agree on an outwards movement of the S4 segment upon activating voltage, but the real-time conformational transitions are still unattainable. With purified human voltage-gated proton (hHv1) channels reconstituted in liposomes, we have examined its conformational dynamics, including the S4 segment at different voltage and pHs using single-molecule fluorescence resonance energy transfer (smFRET). Here, we provide the first glimpse of real-time conformational trajectories of the hHv1 voltage sensor and show that both voltage and pH gradient shift the conformational dynamics of the S4 segment to control channel gating. Our results indicate that the S4 segment transits among 3 major conformational states and kinetic analysis suggest that only the transitions between the inward and outward conformations are highly dependent on voltage and pH changes. Our smFRET studies uncover the stochastic conformational dynamics of S4 and demonstrate how voltage and pH shift its conformational distributions to regulate channel gating. Altogether, we propose a kinetic model that explains the mechanisms underlying voltage and pH gating in Hv channels, which may also serve as a general framework for understanding the voltage sensing and gating in other voltage-gated ion channels.
Data availability
The source data of all smFRET traces, contour maps, histograms, as well as liposome flux assay data is deposited in Dryad (Dryad Digital Repository, doi:10.5061/dryad.dv41ns1zs), including Fig 1b, c, d, e, g, h; Fig 2; Fig 3a and b; Fig 4a, b; Fig 5a, Fig 1-figure supplement1a and c, Fig1-figure supplement 2a, b, c; Fig1-figure supplement 3a, b; Fig1-figure supplement 4a and b.
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Data from: Structural dynamics determine voltage and pH gating in human voltage-gated proton channelDryad Digital Repository, doi:10.5061/dryad.dv41ns1zs.
Article and author information
Author details
Funding
NIH (1R15GM137215-01)
- Shizhen Wang
University of Missouri-Kansas City (Startup fund)
- Shizhen Wang
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2022, Han et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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