Mutation of Asn(111) in the third transmembrane domain of the AT(1A) angiotensin II receptor induces its constitutive activation

A preliminary model of the rat AT(1A) angiotensin II (AII) receptor (Joseph, M. P., Maigret, B., Bonnafous J.-C., Marie, J., and Scheraga, H. A. (1995) J. Protein Chem. 14, 381-398) has predicted an interaction between Asn(111) located in transmembrane domain (TM) In: and Tyr(292) (TM VII) in the nonactivated receptor; a disruption of this interaction upon AII activation would allow Tyr(292) to interact with the conserved Asp(74) (TM II), The previous verification that Ty(292) is essential for receptor coupling to phospholipase C (Marie, J., Maigret, B., Joseph, M. P., Larguier, R., Nouet, S., Lombard, C., and Bonnafous, J.-C. (1994) J. Biol. Chem. 269, 20815-20818) prompted us to check the possible alterations in receptor properties upon Asn(111)-->Ala mutation. The mutated receptor (N111A) displayed: (i) strong constitutive activity, with amplification of the maximal phospholipase C response to AII; (ii) agonist behavior of the AT(2)-specific ligand CGP 42112A, [Sar(1),Ile(8)]AII, and [Sar(1),Ala(8)]AII, antagonists of the wild-type receptor; (iii) inverse agonism behavior of the non-peptide ligands DuP 753, LF 7-0156, and LF 8-0129. The results are discussed in the light of the allosteric ternary complex models and other described examples of constitutive activation of G protein-coupled receptors.