Polynuclear Ruthenium Amines Inhibit K2P Channels via a "Finger in the Dam" Mechanism

Abstract

The trinuclear ruthenium amine ruthenium red (RuR) inhibits diverse ion channels, including K_2P potassium channels, TRPs, the calcium uniporter, CALHMs, ryanodine receptors, and Piezos. Despite this extraordinary array, there is limited information for how RuR engages targets. Here, using X-ray crystallographic and electrophysiological studies of an RuR-sensitive K_2P, K_2P2.1 (TREK-1) I110D, we show that RuR acts by binding an acidic residue pair comprising the "Keystone inhibitor site" under the K_2P CAP domain archway above the channel pore. We further establish that Ru360, a dinuclear ruthenium amine not known to affect K_2Ps, inhibits RuR-sensitive K_2Ps using the same mechanism. Structural knowledge enabled a generalizable design strategy for creating K_2P RuR "super-responders" having nanomolar sensitivity. Together, the data define a "finger in the dam" inhibition mechanism acting at a novel K_2P inhibitor binding site. These findings highlight the polysite nature of K_2P pharmacology and provide a new framework for K_2P inhibitor development.

Keywords: CAP domain; K2P channel; Keystone inhibitor site; Ru360; X-ray crystallography; electrophysiology; ruthenium red.