BALANCED AFFINITY AT(1) AT(2) RECEPTOR NONPEPTIDE BINDING-SITE DETERMINANTS ON THE AT(1) ANGIOTENSIN RECEPTOR/

Angiotensin II (Ang II), a potent modulator of cardiovascular homeosta sis, mediates its effects through two Ang II receptor subtypes (AT(1) and AT(2)) which share less than 30% amino acid homology and can be di fferentiated by specific nonpeptide ligands. We investigated the struc tural determinants of ligand binding to the AT(1) receptor for L-163,0 17 which belongs to a newly developed class of nonpeptides that have e quivalent affinities for AT(1) and AT(2) receptors. Nonpeptide binding affinities were determined in radioreceptor binding assays in membran es from COS cells transfected with wild-type and mutant receptor cDNAs . The amphibian AT receptor variant, xCM46, recognized L-163,017 with an affinity (IC50 = 8 +/- 2 nM) that was only 1.5-fold lower than for the rat AT, receptor (IC50 = 5 +/- 1 nM) which is in striking contrast to the poor affinity of the amphibian xAT(a) wild-type receptor (IC50 > 50 mu M). Analysis of single point rAT(1) receptor mutants in which individual mammalian residues were replaced by the corresponding frog amino acids revealed significant overlap but also distinct difference s in ligand binding interactions between dual receptor and subtype sel ective nonpeptides. These data suggest that AT(1)-selective and dual r eceptor nonpeptides share overlapping but distinct binding pockets on the AT(1) receptor. These findings may lead to improved therapeutics f or the treatment of cardiovascular diseases involving both AT receptor subtypes.