Abstract
Voltage-gated ion channels feature voltage sensor domains (VSDs) that exist in three distinct conformations during activation: resting, intermediate, and activated. Experimental determination of the structure of a potassium channel VSD in the intermediate state has previously proven elusive. Here, we report and validate the experimental three-dimensional structure of the human KCNQ1 voltage-gated potassium channel VSD in the intermediate state. We also used mutagenesis and electrophysiology in Xenopus laevisoocytes to functionally map the determinants of S4 helix motion during voltage-dependent transition from the intermediate to the activated state. Finally, the physiological relevance of the intermediate state KCNQ1 conductance is demonstrated using voltage-clamp fluorometry. This work illuminates the structure of the VSD intermediate state and demonstrates that intermediate state conductivity contributes to the unusual versatility of KCNQ1, which can function either as the slow delayed rectifier current (IKs) of the cardiac action potential or as a constitutively active epithelial leak current.
Highlights
Voltage-gated potassium (KV) channels are critical for electrical signaling in excitable cells where they drive action potential termination
It has long been known that voltage sensor domains fold autonomously, as reflected by the fact that voltage-gated proton channels are single domain monomeric VSDs (DeCoursey et al, 2016; Ramsey et al, 2006; Sasaki et al, 2006) and by studies showing that VSDs excised from KV channels or other voltage-regulated proteins fold independently and yield experimental 3D structures that are consistent with their conformations in the context of intact channels (Li et al, 2014b; Jiang et al, 2003)
We conducted the studies of this work in LMPG rather than LPPG micelles because a recent study indicated that the wild type KCNQ1 VSD adopts a stable fold in this medium (Huang et al, 2018)
Summary
Voltage-gated potassium (KV) channels are critical for electrical signaling in excitable cells where they drive action potential termination. Following this line of logic, even if high-resolution VSD structures in the intermediate state were to be determined, the lack of straightforward functional electrophysiology tests to discriminate between VSD conformations of non-conducting channel states (e.g. resting state vs intermediate state) presents a challenge for functional validation In this regard, it is significant that the VSD of the KCNQ1 KV channel is thought to populate an intermediate state that promotes a conductive state of the pore domain (Zaydman et al, 2014; Hou et al, 2017; Hou et al, 2019; Hou et al, 2020), providing a pathway to functional validation of a VSD structure proposed to represent the intermediate state. We provide evidence to demonstrate that the conductive intermediate state of the KCNQ1 channel is physiologically relevant and contributes to the channel’s functional versatility
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