Regions of the alpha 1-adrenergic receptor involved in coupling to phosphatidylinositol hydrolysis and enhanced sensitivity of biological function.

Abstract

Regions of the hamster .alpha.1-adrenergic receptor (.alpha.1AR) that are important in GTP-binding protein (G protein)-mediated activation of phospholipase C were determined by studying the biological functions of mutant receptors constructed by recombinant DNA techniques. A chimeric receptor consisting of the .beta.2-adrenergic receptor (.beta.2AR) into which the putative third cytoplasmic loop of the .alpha.1AR had been placed activated phosphatidylinositol metabolism as effectively as the native .alpha.1AR, as did a truncated .alpha.1AR lacking the last 47 residues in its cytoplasmic tail. Substitutions of .beta.2AR amino acid sequence in the intermediate portions of the third cytoplasmic loop of the .alpha.1AR or the N-terminal portion of the cytoplasmic tail caused marked decreases in receptor coupling to phospholipase C. Conservative substitutions of two residues in the C terminus of the third cytoplasmic loop (Ala293 .fwdarw. Leu, Lys290 .fwdarw. His) increased the potency of agonists for stimulating phosphatidylinositol metabolism by up to 2 orders of magnitude. These data indicate (i) that the regions of the .alpha.1AR that determine coupling to phosphatidylinositol metabolism are similar to those previously shown to be involved in coupling of .beta.2AR to adenylate cyclase stimulation and (ii) that point mutations of a G-protein-coupled receptor can cause remarkable increases in sensitivity of biological response.