IDENTIFICATION OF A 38-KDA HIGH-AFFINITY SULFONYLUREA-BINDING PEPTIDEIN INSULIN-SECRETING CELLS AND CEREBRAL-CORTEX

Previous studies have described specific photoincorporation of radiola beled sulfonylureas into a peptide with high molecular mass (140-175 k Da), which thus has been suggested to represent the sulfonylurea recep tor. In the present study, a I-125-labeled 4-azidosalicyloyl analog of glibenclamide, I-125-N-3-GA (N-[4-(2-(4-azido-2-hydroxy-5- nzamido)et hyl)benzenesulfonyl]-N'-cyclohexylurea), was used for photoaffinity la beling. This novel probe was specifically photoincorporated into a pep tide with an apparent molecular mass of 160-175 kDa when samples from insulin-secreting HIT cells or cerebral cortex were boiled in a SDS-bu ffer prior to separation with SDS-polyacrylamide gel electrophoresis, However, omitting the heating step revealed specific labeling of an ad ditional 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- nzamido)ethyl)benzenesulfonyl]-N'-c yclohexylurea), glibenclamide, glipizide, and tolbutamide. Analysis of photoaffinity labeling of solubilized fractions indicated an almost e xclusive specific linkage to the 38-kDa peptide. The data support the view that the sulfonylurea receptor in insulin-secreting cells and cer ebral 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.