RT Journal Article SR Electronic T1 Characterization of a Mutant Sulfonylurea Receptor SUR2B with High Affinity for Sulfonylureas and Openers: Differences in the Coupling to Kir6.x Subtypes JF Molecular Pharmacology JO Mol Pharmacol FD American Society for Pharmacology and Experimental Therapeutics SP 190 OP 199 DO 10.1124/mol.60.1.190 VO 60 IS 1 A1 Annette Hambrock A1 Cornelia Löffler-Walz A1 Ulrich Russ A1 Ulf Lange A1 Ulrich Quast YR 2001 UL http://molpharm.aspetjournals.org/content/60/1/190.abstract AB ATP-dependent K+ channels are composed of pore-forming subunits of the Kir6.x family and of sulfonylurea receptors (SURs). SUR1, expressed in pancreatic β-cells, has a higher affinity for sulfonylureas, such as glibenclamide, than SUR2B, expressed in smooth muscle. This difference is mainly caused by serine 1237 in SUR1 corresponding to tyrosine 1206 in SUR2B. To increase the affinity of SUR2B for glibenclamide, the mutant SUR2B(Y1206S) was constructed. In whole-cell patch-clamp experiments, glibenclamide inhibited the channel formed by coexpression of mutant SUR2B with Kir6.1 or 6.2 in human embryonic kidney cells with IC50values of 2.7 and 13 nM, respectively (wild-type, 43 and 167 nM). In intact cells, [3H]glibenclamide bound to mutant SUR2B with a K D value of 4.7 nM (wild-type, 32 nM); coexpression with Kir6.1 or 6.2 increased affinity by 4- and 8-fold, respectively. Binding of the opener [3H]P1075 to SUR2B(Y1206S) was the same as to wild-type and was unaffected by coexpression. In cells, the ratio of glibenclamide:P1075 sites was ≈ 1:1; in membranes, it varied with the MgATP concentration. Heterologous competition curves were generally biphasic; the shape of the curve depended on the Kir-subtype. The effects of coexpression were weakened or abolished when binding assays were conducted in membranes. It is concluded that the mutation Y1206S increases the affinity of SUR2B for and the channel sensitivity toward glibenclamide by 7- to 15-fold. The interaction of glibenclamide (but not opener) with mutant SUR2B is modified by coexpression with Kir6.x in a manner depending on the Kir subtype and on the integrity of the cell. KATP channelATP-sensitive K+ channelSUsulfonylureaSURsulfonylurea receptorP1075N-cyano-N′-(1,1-dimethylpropyl)-N"-3-pyridylguanidineKirinwardly rectifying K+ channelHEKhuman embryonic kidneyGBCglibenclamideBNSnonspecific bindingBTOTtotal bindingBmaxmaximum concentration of binding sitesBSspecific bindingAZ-DF 2654-[[N-(α-phenyl-2-piperidino-benzyl)carbamoyl]methyl]benzoic acidPIP2phosphatidylinositol-4,5-bisphosphate