Structural basis for K2P channel gating and modulation

PROJECT SUMMARY
This project aims to understand the function and regulation of three ion channels that conduct K+ across cell
membranes and belong to the two-pore domain K+ channel family. We will apply cryo-electron microscopy to
determine structures of the channels in different functional states within lipid environments that mimic the
cellular membrane and electrophysiological recordings to characterize their activities in order to derive physical
models for channel gating and modulation. We aim to capture structural snapshots of the different open and
closed conformations for each channel by varying conditions that alter channel function including solution
composition, lipid composition, and presence of small molecules or interacting proteins. The three channels
are members of different branches of the two-pore domain K+ channel family. While they share a common
structural architecture, each channel is regulated by pH in a different way; one is inhibited by protons on both
sides of the membrane, the second is inhibited only by extracellular protons, and the third is both inhibited and
altered in its ionic selectivity by extracellular protons. The underlying molecular mechanisms by which pH is
sensed and converted into a change in channel activity are correspondingly different between the channels.
Each channel is further regulated by a distinct set of factors including signaling lipids, interacting proteins,
solution ion composition, and small molecule drugs. Comparative analyses of the three structurally and
evolutionarily related K+ channels will therefore provide additional insight into their functional properties and
biological roles. Two-pore domain K+ channels mediate cellular electrical signaling by establishing and
maintaining the resting membrane potential and opposing excitability. The channels under study here are
involved in respiratory regulation, cardiac rhythm generation, blood pressure control, central chemoreception,
and systemic pH homeostasis among other processes. Their dysregulation is implicated in cardiac arrythmia,
kidney disease, and hypertension in humans and they are targets of anesthetics, antiarrhythmics, and drugs
under investigation for obstructive sleep apnea. Therefore, in addition to providing fundamental mechanistic
insight into the physical and chemical basis for channel function, this work will serve as a basis for the
development of more potent and specific pharmacological agents targeting ion channels to promote health and
treat disease. Importantly, the technical and methodological advances developed here for structural
characterization of small membrane proteins in lipid environments are expected to be widely applicable and will
facilitate insights across the breadth of biology in which membrane proteins play important roles.

Grant Number: 5R01GM145869-04
NIH Institute/Center: NIH
Principal Investigator: Stephen Brohawn