title: pH-dependent gating of ROMK (Kir1.1) channels involves conformational changes in both N and C termini.
abstract: ROMK channels (Kir1.1) are members of the superfamily of inward rectifier potassium channels (Kir) and represent the channels underlying K+ secretion in the kidney. As their native counterparts, Kir1.1 channels are gated by intracellular pH, with acidification leading to channel closure. Although a lysine residue (Lys80) close to the first hydrophobic segment M1 has been identified as the pH sensor, little is known about how opening and closing of the channel is accomplished. Here we investigate the gating process of Kir1.1 channels exploiting their state-dependent modification by water-soluble oxidants and sulfhydryl reagents. Mutagenesis of all intracellular cysteines either alone or in combination revealed two residues targeted by these reagents, one in the N terminus (Cys49) and one in the C terminus (Cys308) of the channel protein. Both sites reacted with the thiol reagents only in the closed state and not in the open state. These results indicate that pH-dependent gating of Kir1.1 channels involves movement of protein domains in both N and C termini of the Kir1.1 protein.
authors.affiliation: Department of Physiology II, Ob dem Himmelreich 7, 72074 Tübingen, Germany.
authors.forename: U
authors.initials: U
authors.lastname: Schulte
authors.forename: H
authors.initials: H
authors.lastname: Hahn
authors.forename: H
authors.initials: H
authors.lastname: Wiesinger
authors.forename: J P
authors.initials: JP
authors.lastname: Ruppersberg
authors.forename: B
authors.initials: B
authors.lastname: Fakler
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D003545:Cysteine; D004228:Dithionitrobenzoic Acid / Q000494:pharmacology; D004229:Dithiothreitol / Q000494:pharmacology; D005260:Female; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D015640:Ion Channel Gating / Q000187:drug effects / Q000502:physiology*; D007668:Kidney / Q000502:physiology; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D016297:Mutagenesis, Site-Directed; D009865:Oocytes / Q000502:physiology; D010084:Oxidation-Reduction; D018408:Patch-Clamp Techniques; D015221:Potassium Channels / Q000737:chemistry / Q000502:physiology*; D024661:Potassium Channels, Inwardly Rectifying; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D014982:Xenopus laevis
chemical_list: D015221:Potassium Channels; D024661:Potassium Channels, Inwardly Rectifying; D011994:Recombinant Proteins; D004228:Dithionitrobenzoic Acid; D003545:Cysteine; D004229:Dithiothreitol