Identification of two serine residues involved in agonist activation of the beta-adrenergic receptor.

Pharmacophore mapping of the ligand binding domain of the beta-adrenergic receptor has revealed specific molecular interactions which are important for agonist and antagonist binding to the receptor. Previous site-directed mutagenesis experiments have demonstrated that the binding of amine agonists and antagonists to the receptor involves an interaction between the amine group of the ligand and the carboxylate side chain of Asp113 in the third hydrophobic domain of the receptor (Strader, C. D., Sigal, I. S., Candelore, M. R., Rands, E., Hill, W. S., and Dixon, R. A. F. (1988) J. Biol. Chem. 263, 10267-10271). We have now identified 2 serine residues, at positions 204 and 207 in the fifth hydrophobic domain of the beta-adrenergic receptor, which are critical for agonist binding and activation of the receptor. These serine residues are conserved with G-protein-coupled receptors which bind catecholamine agonists, but not with receptors whose endogenous ligands do not have the catechol moiety. Removal of the hydroxyl side chain from either Ser204 or Ser207 by substitution of the serine residue with an alanine attenuates the activity of catecholamine agonists at the receptor. The effects of these mutations on agonist activity are mimicked selectively by the removal of the catechol hydroxyl moieties from the aromatic ring of the agonist. The data suggest that the interaction of catecholamine agonists with the beta-adrenergic receptor involves two hydrogen bonds, one between the hydroxyl side chain of Ser204 and the meta-hydroxyl group of the ligand and a second between the hydroxyl side chain of Ser207 and the para-hydroxyl group of the ligand.