Role of Charged Residues in the Regulation of Voltage Sensor Movement in Herg K+ Channels

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The human Ether-a-go-go Related Gene (hERG) K+ channel plays a central role in cardiac action potential repolarization. Compared to other Kv channels where fast activation is due to rapid voltage sensor (S4) movement, hERG activation is slow and the mechanisms remain under debate. Previous studies have shown that charges in the voltage sensing domain contribute to determining the rate of channel activation, which includes transition of the S4 and downstream events in S4-pore coupling. The functional role of individual charges in S4 movement alone is still unclear in hERG channels. We have investigated the effect of neutralization of positive charges (K525-K538) on the voltage dependence of S4 equilibrium by examining the voltage and time-dependent accessibility of cysteine substituted I521 in the tip of the S4. At −120 mV, 521C is almost inaccessible to MTSET for R531Q, R537Q and K538Q, but is accessible for R528Q, R534Q and K525Q. This suggests that at −120 mV, position 521 in the R531Q and R537Q, K538Q mutants is mostly buried, whereas it is extruded in the other mutants. The slopes of the S4 equilibrium curves of K538Q and R531Q are greatly altered, indicating that K538Q favors the transition of S4 from the resting to activated states, while R531Q favors the reverse transitions. To extract the kinetics of S4 movement, we examined the dependence of the MTSET modification rate on pulse duration. We found that time constants of S4 equilibrium for R528:I521C, R537:I521C and R531Q:I521C are slower than control, with the largest effect seen for R531Q:I521C. Our data suggest that K525, R528, R534 normally play a role in stabilizing S4 in the resting state; while R528, R537 and R531 facilitate the transition of S4 from the resting to the activated state.