title: Molecular mechanism of a COOH-terminal gating determinant in the ROMK channel revealed by a Bartter's disease mutation.
abstract: The ROMK subtypes of inward-rectifier K(+) channels mediate potassium secretion and regulate NaCl reabsorption in the kidney. Loss-of-function mutations in this pH-sensitive K(+) channel cause Bartter's disease, a familial salt wasting nephropathy. One disease-causing mutation truncates the extreme COOH-terminus and induces a closed gating conformation. Here we identify a region within the deleted domain that plays an important role in pH-dependent gating. The domain contains a structural element that functionally interacts with the pH sensor in the cytoplasmic NH(2)-terminus to set a physiological range of pH sensitivity. Removal of the domain shifts the pK(a) towards alkaline pH values, causing channel inactivation under physiological conditions. Suppressor mutations within the pH sensor rescued channel gating and trans addition of the cognate peptide restored pH sensitivity. A specific interdomain interaction was revealed in an in vitro protein-protein binding assay between the NH(2)- and COOH-terminal cytoplasmic domains expressed as bacterial fusion proteins. These results provide new insights into the molecular mechanisms underlying Kir channel regulation and channel gating defects that are associated with Bartter's disease.
authors.affiliation: Department of Physiology, University of Maryland School of Medicine, 655 W. Baltimore Street, Baltimore, MD 21201, USA.
authors.forename: Thomas P
authors.initials: TP
authors.lastname: Flagg
authors.forename: Dana
authors.initials: D
authors.lastname: Yoo
authors.forename: Christopher M
authors.initials: CM
authors.lastname: Sciortino
authors.forename: Margaret
authors.initials: M
authors.lastname: Tate
authors.forename: Michael F
authors.initials: MF
authors.lastname: Romero
authors.forename: Paul A
authors.initials: PA
authors.lastname: Welling
mesh_terms: D000595:Amino Acid Sequence / Q000502:physiology; D000818:Animals; D001477:Bartter Syndrome / Q000235:genetics*; D003593:Cytoplasm / Q000378:metabolism; D005260:Female; D016153:Genes, Suppressor / Q000502:physiology; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007425:Intracellular Membranes / Q000378:metabolism; D015640:Ion Channel Gating / Q000502:physiology*; D009154:Mutation; D009865:Oocytes; D015221:Potassium Channels / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D024661:Potassium Channels, Inwardly Rectifying; D017434:Protein Structure, Tertiary / Q000502:physiology; D012016:Reference Values; D014982:Xenopus laevis
chemical_list: C086989:KCNJ1 protein, human; D015221:Potassium Channels; D024661:Potassium Channels, Inwardly Rectifying