HERG Activation Gating Is Rapid: Evidence from Gating-Current Recordings and MTSET Modification of the Voltage Sensor

Abstract

Background The human ether-a-go-go related gene protein (hERG) K + channel (Kv11.1) plays a major role in cardiac action potential repolarization, and defects in its function/expression can lead to long QT syndrome. The channel's key physiologic function as the main repolarization current is attributable in part to its unconventional slow activation kinetics, which are an order of magnitude slower than other members of the voltage-gated potassium channel superfamily. Methods We recorded gating-currents of hERG channels in tsa201 cells in order to directly identify the rate-limiting step in the activation of hERG channels. Expression of hERG channels was optimized by subcloning channel cDNA into a pGW1H expression vector and transfection into tsa201 cells. Gating currents were recorded in NMG + -based potassium-free internal and external solutions. As well, we used MTS modification of an externally facing residue in S4, I521C, to track the time course of movement of the S4 voltage sensor. I521 was mutated to a cysteine residue expressed in Xenopus or tsa201 cells, and various thiol-modifying reagents were used to modulate hERG activation and deactivation kinetics. Results hERG gating currents recorded from tsa201 cells were observed to activate on a millisecond time scale with time constants of on-gating current movement at 0 mV of 3.7 and 11.3 ms, much faster than ionic current activation, with bimodal voltage dependence (V 1/2-1 = –36.6 mV, k 1 = 8.3 mV, V 1/2-2 = 42.8 mV, k 2 = 15.5 mV, r² = 0.9912; A1=26%). Direct outward movement of the S4 domain as tracked by the modification of hERG channel I521C by MTSET was complete at 50 ms for pulses to 0 mV and within 25 ms for pulses to +60 mV. Conclusions This study represents the first report of hERG gating currents observed from mammalian cells and the first report of rapid voltage sensor modification associated with activation in hERG channels. Gating current recordings from tsa201 cells and the rate of MTSET modification of hERG voltage sensors suggest that the delayed activation kinetics of hERG cannot be attributed to slow voltage sensor movement but may be rate limited by a slow coupling process to the opening pore.