Identification of a 38-kDa high affinity sulfonylurea-binding peptide in insulin-secreting cells and cerebral cortex.
Previous studies have described specific photoincorporation of radiolabeled sulfonylureas into a peptide with high molecular mass (140-175 kDa), which thus has been suggested to represent the sulfonylurea receptor. In the present study, a 125I-labeled 4-azidosalicyloyl analog of glibenclamide, 125I-N3-GA (N-[4-(2-(4-azido-2-hydroxy-5-125I- iodobenzamido)ethyl)benzenesulfonyl]-N'-cyclohexylurea), was used for photoaffinity labeling. This novel probe was specifically photoincorporated into a peptide with an apparent molecular mass of 160-175 kDa when samples from insulin-secreting HIT cells or cerebral cortex were boiled in a SDS-buffer prior to separation with SDS-polyacrylamide gel electrophoresis. However, omitting the heating step revealed specific labeling of an additional peptide with an apparent molecular mass of 38 kDa. The amount of radioactivity specifically photoincorporated into this peptide was 3-4-fold higher than that incorporated into the 160-175-kDa peptide. Both peptides displayed similar dissociation constants for binding of the sulfonylureas IN3-GA (N-[4-(2-(4-azido-2-hydroxy- 5-iodobenzamido)ethyl)benzenesulfonyl]-N'-cyclohexylurea), glibenclamide, glipizide, and tolbutamide. Analysis of photoaffinity labeling of solubilized fractions indicated an almost exclusive specific linkage to the 38-kDa peptide. The data support the view that the sulfonylurea receptor in insulin-secreting cells and cerebral cortex consists of a peptide with an apparent molecular mass of 38 kDa, which seems to be tightly coupled to a 160-175-kDa peptide.
Institute of Pharmacology and Toxicology, University of Göttingen, Federal Republic of Germany.
M
M
Schwanstecher
S
S
Löser
F
F
Chudziak
U
U
Panten
D018528:ATP-Binding Cassette Transporters; D000345:Affinity Labels; D000818:Animals; D002460:Cell Line; D002540:Cerebral Cortex / Q000378:metabolism*; D007328:Insulin / Q000378:metabolism; D000078790:Insulin Secretion; D007515:Islets of Langerhans / Q000378:metabolism; D015221:Potassium Channels / Q000032:analysis*; D024661:Potassium Channels, Inwardly Rectifying; D011955:Receptors, Drug / Q000032:analysis*; D013453:Sulfonylurea Compounds / Q000378:metabolism*; D064233:Sulfonylurea Receptors; D013552:Swine
D018528:ATP-Binding Cassette Transporters; D000345:Affinity Labels; D007328:Insulin; D015221:Potassium Channels; D024661:Potassium Channels, Inwardly Rectifying; D011955:Receptors, Drug; D013453:Sulfonylurea Compounds; D064233:Sulfonylurea Receptors