External Protons Destabilize the Relaxed State of the hERG Channel Voltage Sensor

Abstract

Voltage sensor relaxation may contribute to the slow deactivation kinetics of hERG channels1. Relaxation stabilizes the voltage sensor in the activated state resulting in a mode-shift, where the voltage-dependence of channel deactivation is shifted ∼-30 mV compared with that of channel activation. Voltage clamp fluorimetry (VCF) reports of voltage sensor movement show a similar shift in the voltage-dependence of S4 return compared with its activation. However, the mechanism underlying voltage sensor relaxation in hERG is unknown. In Shaker channels, voltage sensor relaxation has been shown to be sensitive to the length and composition of the S3-S4 linker2. Here, we assessed the role of S3-S4 linker length and composition in voltage sensor relaxation in hERG channels using two constructs: a chimera with the 9 residue hERG S3-S4 linker substituted with the 31 residue Shaker S3-S4 linker; the other with the 9 residues of the hERG S3-S4 linker mutated to glycine residues. In both cases, the robust shifts in the voltage-dependence of ionic current deactivation relative to that of activation were preserved (−35 and −29 mV in the 31 residue Shaker linker construct (n=4) and the 9 glycine construct (n=5), respectively), suggesting that length and composition of the S3-S4 linker are not important mediators of hERG channel relaxation. Instead, we demonstrate that the ionic current mode-shift is reduced by external protons in a pH-dependent manner with a pKa of 5.5 (n=5). Furthermore, VCF recordings of voltage sensor movement show that acidic pH directly inhibits voltage sensor relaxation. These data suggest that external protons modify voltage sensor relaxation by destabilizing the relaxed conformation and reveal a mechanism by which acidic pH may accelerate hERG channel deactivation gating kinetics. 1.Tan et al., J.Gen.Physiol. 140:293-306, 2012 2.Priest et al., Biophys.J. 105:2312-2322, 2013