GLUCAGON-CENTER-DOT-GLUCAGON-LIKE PEPTIDE-I RECEPTOR CHIMERAS REVEAL DOMAINS THAT DETERMINE SPECIFICITY OF GLUCAGON BINDING

The binding of glucagon to its hepatic receptor triggers a G-protein-m ediated signal that ultimately leads to an increase in hepatic glucose production (gluconeogenesis) and glycogen breakdown (glycogenolysis). In order to elucidate the structural domain(s) of the human glucagon receptor (hGR) involved in the selective binding of glucagon, a series of chimeras was constructed in which various domains of the hGR were replaced by homologous regions from the receptor for the glucoin-creti n hormone, glucagon-like peptide I (GLP-IR), hGR and GLP-IR are quite similar (47% amino acid identity) yet have readily distinguishable lig and binding characteristics; glucagon binds to the recombinant hGR exp ressed in COS-7 cells with a K-d that is 1000-fold lower than the K-d for glucagon binding to GLP-IR, In the present study, chimeric recepto rs were transiently expressed in COS-7 cells and analyzed for glucagon binding, Expression of each receptor chimera was confirmed by immunof luorescence staining using a hGR-specific monoclonal antibody, This re port identifies several noncontiguous domains of the hGR that are impo rtant for high affinity glucagon binding, Mast notable are the membran e-proximal half of the amino-terminal extension, the first extracellul ar loop, and the third, fourth, and sixth transmembrane domains.