TYR(292) IN THE 7TH TRANSMEMBRANE DOMAIN OF THE AT(1A) ANGIOTENSIN-IIRECEPTOR IS ESSENTIAL FOR ITS COUPLING TO PHOSPHOLIPASE-C

An essential role of the conserved Asp(74) in, the coupling of the typ e 1 angiotensin II (AII) receptor (AT(1)) to phospholipase C has alrea dy been reported (Bihoreau, C., Monnot, C., Davies, E., Teutsch, B., B ernstein, K. B., Corvol, P., and Clauser, E. (1993) Proc. Natl. Acad. Sci. U.S. A. 90, 5133-5137). Moreover, preliminary modeling studies ha ve shown that a spatial proximity exists between Asp(74), located in t ransmembrane domain II, and Tyr(292), located in transmembrane domain VII and conserved in many, but not all, G protein-coupled receptors. W e mutated Tyr(292) into Phe and evaluated the pharmacological and acti vation characteristics of the mutated receptor (Y292F) stably expresse d in Chinese hamster ovary cells. This receptor possessed unchanged bi nding properties for agonist or antagonist peptide ligands compared to the wild-type receptor, while its coupling to phospholipase C was sev erely impaired. Interestingly, competition binding experiments, using I-125-[Sar(1)]AII as a tracer ligand, showed that the Y292F receptor d isplayed an increased K-i value for DuP 753, an AT(1)-specific nonpept ide antagonist and a greatly decreased K-i value for the AT(2)-specifi c ligand CGP 42112A. These pharmacological changes are similar to thos e observed for the previously reported mutation of Asp(74) into Asn. T his apparently symmetrical role of Asp(74) and Tyr(292) is consistent with the hypothesis that an interaction between these two amino acids could be a key event in the molecular processes linking AII recognitio n and AT(1) receptor activation.