POLAR RESIDUES IN THE TRANSMEMBRANE DOMAINS OF THE TYPE-1 ANGIOTENSIN-II RECEPTOR ARE REQUIRED FOR BINDING AND COUPLING - RECONSTITUTION OFTHE BINDING-SITE BY COEXPRESSION OF 2 DEFICIENT MUTANTS

Type 1 angitensin receptors (AT(1)) are G protein coupled receptors, m ediating the physiological actions of the vasoactive peptide ansotensi n II, In this study, the roles of 7 amino acids of the rat AT(1A) rece ptor in ligand binding and signaling were investigated by performing f unctional assays of individual receptor mutants expressed in COS and C hinese hamster ovary cells, Substitutions of polar residues in the thi rd transmembrane domain with Ala indicate that Ser(105), Ser(107), and Ser(109) are not essential for maintenance of the ansotensin II bindi ng site, Replacement of Asn(111) or Ser(115) does not alter the bindin g affinity for peptidic analogs, but modifies the ability of the recep tor to interact with AT, (DuP753)- or AT(2) (CGP42112A)-specific ligan ds, These 2 residues are probably involved in determining the binding specificity for these analogs, The absence of G-protein coupling to th e Ser(115) mutant suggests that this residue, in addition to previousl y identified residues, Asp(74) and Tyr(292), participates in the recep tor activation mechanism. Finally, Lys(102) (third helix) and Lys(199) (fifth helix) mutants do not bind angiotensin II or different analogs , Go-expression of these two deficient receptors permitted the restora tion of a normal binding site, This effect was not due to homologous r ecombination of the cDNAs but to protein trans complementation.