Spannungs- und pH-Steuerung des Hv1 Protonenkanals
Zusammenfassung der Projektergebnisse
The Hv1 proton channel is gated by both voltage and the pH difference across the membrane (delta pH); the mechanism of this coupling is not well understood. We identified Delta pH-induced conformational changes at the S4 voltage sensor of the "Ciona intestinalis" proton channel (ciHv1) with patch-clamp fluorometry and the substituted cysteine-accessibility method. We could show that during the resting state of Hv1, at negative membrane potentials, the S4 sensor can change its conformation in response to changes in delta pH. Acidification at one side of the membrane push the S4 sensor to the other side of the membrane. We confirmed these results by using caged protons to rapidly change delta pH by light-induced release of protons during patch-clamp fluorometry recordings. Changes in delta pH do not uncouple gating from motion of the S1 segment, suggesting that the S1 segment is not directly involved in delta pH sensing. We identified amino-acid positions at the extracellular side of the S4 voltage sensor facing the lumen of the VSD, isoleucine 248 and leucine 251, that contribute to delta pH-sensing: while the conductance-voltage (GV) relationship of wildtype ciHv1 shifts by ~45 mV/delta pH unit, the GV relationships of ciHv1 mutated at those positions shift by only around 22 mV/delta pH unit. We suggest that S4 functions as a dual sensor, which integrates changes in voltage and delta pH. Both signals position S4 in the membrane and codetermine the opening probability. In addition, we developed photo-controllable blockers of Hv1 by synthetizing azobenzene-based guanidine derivatives and proved their efficiency to block proton currents in human sperm and macrophages. These blockers allow the investigation of the physiological role of Hv1 with high spatiotemporal precision. Finally, we identified and characterized a second voltage-gated and proton-selective channel, HCNL1. Physiologically, HCNL1 has the opposite role of Hv1: HCNL1 activation leads to a proton influx, while Hv1 activation leads to a proton efflux. HCNL1, akin to Hv1, also conducts protons via the voltage-sensor domain. However, HCNL1 forms a tetramer and belongs to the family of hyperpolarization-activated ion channels. Thus, voltage-gated proton channels were invented at least twice during evolution.
Projektbezogene Publikationen (Auswahl)
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Photocontrol of the Hv1 proton channel. (2017) ACS Chem. Biol. 12:2952-7
Rennhack, A, Grahn, E, Kaupp, UB, Berger TK
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Post-translational cleavage of Hv1 in human sperm tunes pH- and voltage-dependent gating. (2017) J. Physiol. 595:1533-46
Berger TK, Fußhöller DM, Goodwin N, Bönigk W, Müller A, Dokani Khesroshahi N, Brenker C, Wachten D, Krause E, Kaupp UB, Strünker T
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A family of hyperpolarization-activated channels selective for protons. (2020) Proc. Natl. Acad. Sci. U S A 117:13783-91
Wobig L, Wolfenstetter T, Fechner S, Bönigk W, Körschen HG, Jikeli JF, Trötschel C, Feederle R, Kaupp UB, Seifert R, Berger TK
