Multisite contacts involved in coupling of the β‐adrenergic receptor with the stimulatory guanine‐nucleotide‐binding regulatory protein: Structural and functional studies by β‐receptor‐site‐specific synthetic peptides

Synthetic peptides, 12–22 amino acid residues long, comprising the presumed coupling sites of the β‐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 μM, or 3–4 μM 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‐adenylate‐cyclase 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 α subunit of G_s and downstream effector regulation. This model corroborates and expands a similar model suggested for activated rhodopsin‐transducin interaction [König, B., Arendt, A., McDowell, J. H., Kahlert, M., Hargrave, P. A. & Hofmann, K. P. (1989) Proc. Natl Acad. Sci. USA 86, 6878–6882.