title: Identification of the high-affinity tolbutamide site on the SUR1 subunit of the K(ATP) channel.
abstract: ATP-sensitive potassium channels (K(ATP)) are formed from four pore-forming Kir6.2 subunits complexed with four regulatory sulfonylurea receptor subunits (SUR1 in pancreatic beta-cells, SUR2A in heart). The sensitivity of the channel to different sulfonylureas depends on the SUR isoform. In particular, Kir6.2-SUR1 but not Kir6.2-SUR2A channels are blocked by tolbutamide with high affinity. We made chimeras between SUR1 and SUR2A to identify the region of the protein involved in high-affinity tolbutamide block. Chimeric SURs were coexpressed with Kir6.2 in Xenopus oocytes, and macroscopic currents were measured in inside-out membrane patches. High-affinity tolbutamide inhibition could be conferred on SUR2A by replacing transmembrane domains (TMs) 14-16 with the corresponding region of SUR1. Conversely, high-affinity tolbutamide inhibition of SUR1 was abolished by replacing TMs 13-16 with the corresponding SUR2A sequence, or by mutating a single serine residue within this region to tyrosine (S1237Y). Binding of [3H]glibenclamide to membranes expressing SUR1 was abolished concomitantly with the loss of high-affinity tolbutamide block. These results suggest that a site in the COOH-terminal set of TMs of the SUR1 subunit of the K(ATP) channel is involved in the binding of tolbutamide and glibenclamide.
authors.affiliation: Nuffield Department of Clinical Biochemistry, John Radcliffe Hospital, Oxford, UK.
authors.forename: R
authors.initials: R
authors.lastname: Ashfield
authors.forename: F M
authors.initials: FM
authors.lastname: Gribble
authors.forename: S J
authors.initials: SJ
authors.lastname: Ashcroft
authors.forename: F M
authors.initials: FM
authors.lastname: Ashcroft
mesh_terms: D018528:ATP-Binding Cassette Transporters; D000244:Adenosine Diphosphate / Q000378:metabolism; D019943:Amino Acid Substitution; D000818:Animals; D001665:Binding Sites; D019556:COS Cells; D005905:Glyburide / Q000378:metabolism; D007004:Hypoglycemic Agents / Q000378:metabolism*; D015221:Potassium Channels / Q000235:genetics / Q000378:metabolism*; D024661:Potassium Channels, Inwardly Rectifying; D051381:Rats; D011955:Receptors, Drug / Q000235:genetics / Q000378:metabolism*; D013453:Sulfonylurea Compounds / Q000378:metabolism*; D064233:Sulfonylurea Receptors; D014044:Tolbutamide / Q000378:metabolism*; D014162:Transfection; D014981:Xenopus
chemical_list: D018528:ATP-Binding Cassette Transporters; C577509:Abcc8 protein, rat; D007004:Hypoglycemic Agents; D015221:Potassium Channels; D024661:Potassium Channels, Inwardly Rectifying; D011955:Receptors, Drug; D013453:Sulfonylurea Compounds; D064233:Sulfonylurea Receptors; D000244:Adenosine Diphosphate; D014044:Tolbutamide; D005905:Glyburide