A Molecular Mechanism for Gating Polarity in Non-Domain-Swapped Kv Channels

Abstract

The molecular basis of ion channel gating polarity, whether a channel opens via depolarization or hyperpolarization, is poorly understood. This is a consequence of multiple factors but can be traced to the inherent difficulties in generalizing channel behaviors between channels sharing low sequence and structural similarity. We address this issue by focusing on the properties that define gating polarity in the plant Kv channels KAT1 and SKOR. These are hyperpolarization (KAT1) and depolarization (SKOR ) activated Kv channels sharing, respectively, 47% identity within their transmembrane domains. Using the recently determined cryo-EM structure of KAT1 as a baseline, we conducted a mutagenic walk between KAT1 and SKOR, monitoring macroscopic currents of the mutants and always referencing the results to the 3D structure. Our results uncover several crucial positions at the intracellular interface between the VSD and the pore, in the S5, S6, and proximal C-linker that shift the gating polarity of KAT1, turning it in an outward rectifying channel. These results are analyzed in the context of a mechanistic model where the interaction between the downwards-moving S4 and the C-linker defines electromechanical coupling in these channels. GFC and MC are equally contributed to this work.