Pore Helix Mediates Proton Block of Vanilloid Receptors

Abstract

Tissue acidosis occurs during inflammation and injury, and modulates many receptors and ion channels on the pain pathway including the capsaicin ion channel TRPV1. Extracellular low pH exerts several effects on the function of TRPV1. Extreme acidification leads to direct activation of the channel, while mild acidic pH potentiates its response to other stimuli. Paradoxically, protons also inhibit the unitary conductance of the channel. This inhibitory effect confers TRPV1 a similar maximum response at low pH in spite of increased agonist sensitivity, thereby limiting the ion flux into cells. Proton-mediated pore block has been studied extensively in other ion channels. Two representative mechanisms have been proposed, one involving competitive inhibition with permeating ions, and the other by reducing the surface potential of membranes. We have examined these mechanisms for proton block of TRPV1. Surprisingly, we found that neither mechanism could adequately account for the full blocking effect of protons. Mutagenesis experiments revealed that, in addition to a residue at the pore entrance, another residue on the pore helix was also responsible, each of which accounted for only half of the total inhibition. Only double mutations of both residues could eliminate the full blockade. In the prototype Kcsa structure, the counter residues have been suggested to form a hydrogen bond, and the disruption of their interaction destabilizes the pore conductance. These results suggest that protons may block TRPV1 by titrating the interaction of the pore helix with the selectivity filter, and thus provides a novel model for proton-mediated pore block of ion channels.