UC Berkeley

Two-pore domain potassium channels (K2Ps) are widely expressed ion channels which generate leak-type currents regulated by diverse chemical and physical stimuli including temperature, membrane tension, signaling lipids, and pH, to control the resting membrane potential (RMP) of cells. While these K2Ps are critical to the maintenance of the RMP, the structural mechanisms by which they respond to physiological stimuli remain largely unknown. Additionally, studies of K2Ps in vivo have been hampered by the lack of molecular tools which can be used to probe K2P function and structure.

Here, we describe the structural basis for gating of the K2P TWIK1 and describe a series of molecular tools generated against the K2P TREK1. Utilizing cryo-electron microscopy (cryo-EM), we determine high resolution structure of TWIK1 in lipid nanodiscs at a high and low pH, and in high extracellular K+, and describe the mechanistic basis for pH-gating in this channel. We then describe the generation of a nanobody which specifically binds another K2P, TREK1, with nanomolar affinity. Finally, we describe the computational prediction of a series of novel small-molecule TREK1 agonists. The data and molecular tools presented here further our understanding of the structural and functional properties of the K2P family of ion channels.