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Molecular mechanism of deactivating gating in HCN2 channels Yip, Delbert


Hyperpolarization-activated Cyclic Nucleotide-gated (HCN) channels open with membrane hyperpolarization, rather than depolarization, unlike most other voltage-gated ion channels. Hyperpolarization-dependent opening in HCN channels is thought to occur by steric inhibition of the pore gate by the voltage-sensing domain (VSD). Recent studies have described molecular movements of voltage sensors upon hyperpolarization and pore opening, along with preliminary opening events in the pore, yet little is known about the open structure of the pore. Thus, the structural mechanism of channel deactivation, and how it differs from activation, is unclear. Here, we analyzed a mutation, F431A, in the distal S6 of mHCN2 channels that greatly slows deactivation over a wide range of voltages, but only modestly slows activation. A four-state allosteric model suggests that F431A impairs voltage-sensor relaxation, but does not alter the steady-state voltage-dependence of activation. The model could also describe voltage hysteresis in WT and F431A channels, but to do so, it required a re-calibration separate from that for isochronal recordings. Our results identify that F431 differentially regulates activation and deactivation gating, and suggest that F431A perturbs interactions between the N-terminal S5 and C-terminal S6, particularly during a voltage-dependent closing step that is late, slow, and rate-limiting.

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