MULTISITE CONTACTS INVOLVED IN COUPLING OF THE BETA-ADRENERGIC-RECEPTOR WITH THE STIMULATORY GUANINE-NUCLEOTIDE-BINDING REGULATORY PROTEIN - STRUCTURAL AND FUNCTIONAL-STUDIES BY BETA-RECEPTOR-SITE-SPECIFIC SYNTHETIC PEPTIDES

Synthetic peptides, 12-22 amino acid residues long, comprising the presumed coupling sites of the beta-adrenergic receptor with the stimulatory guanine-nucleotide-binding regulatory protein (G(s)), were examined for their ability to modulate G(s) activation in turkey erythrocyte membranes. Three peptides corresponding to the second cytoplasmic loop, the N-terminal region of the third cytoplasmic loop, and the N-terminal region of the putative fourth cytoplasmic loop, compete synergistically with the hormone-stimulated receptor for G(s) activation with median effector concentrations of 15-35-mu-M, or 3-4-mu-M for combinations of two peptides. One peptide, corresponding to the C-terminal region of the third cytoplasmic loop, carries the unique ability to activate the G(s)-adenylatecyclase complex independent of the signalling state of the receptor. These observations are consistent with a dynamic model of receptor-mediated G-protein activation in membranes, where domains composed of the second, third and fourth intracellular loop of the receptor bind to and are interactive with the G-protein heterotrimer, resulting in ligand-induced conformational changes of the receptor. In response to hormone binding, the extent or the number of sites involved in interaction with G(s) may be readjusted using a fourth site. Modulation of coupling sites may elicit congruent conformational changes within the G(s) heterotrimer, with qualitatively different effects on GTP/GDP exchange in the alpha-subunit of G(s) and downstream effector regulation. This model corroborates and expands a similar model suggested for activated rhodopsin-transducin interaction.