Subunit-subunit interactions are critical for proton sensitivity of ROMK: Evidence in support of an intermolecular gating mechanism

The tetrameric K channel ROMK provides an important pathway for K secretion by the mammalian kidney, and the gating of this channel is highly sensitive to changes in cytosolic pH. Although charge-charge interactions have been implicated in pH sensing by this K channel tetramer, the molecular mechanism linking pH sensing and the gating of ion channels is poorly understood. The x-ray crystal structure KirBac1.1, a prokaryotic ortholog of ROMK, has suggested that channel gating involves intermolecular interactions of the Wand C-terminal domains of adjacent subunits. Here we studied channel gating behavior to changes in pH using giant patch clamping of Xenopus laevis oocytes expressing WT or mutant ROMK, and we present evidence that no single charged residue provides the pH sensor. Instead, we show that N-C- and C-C-terminal subunit-subunit interactions form salt bridges, which function to stabilize ROMK in the open state and which are modified by protons. We identify a highly conserved C-C-terminal arginine-glutamate (R-E) ion pair that forms an intermolecular salt bridge and responds to changes in proton concentration. Our results support the intermolecular model for pH gating of inward rectifier K channels.