Mechanism of ligand-receptor interaction in yeast

Abstract

A detailed biochemical characterization of the yeast {dollar}\alpha{dollar}-factor receptor was carried out in order to understand the mechanism of ligand/receptor interaction in yeast. The {dollar}\alpha{dollar}-factor receptor is a member of the large family of G protein-coupled, seven transmembrane segment, integral membrane protein receptors. Specificities for ligand binding and activation of cellular responses were separately assayed for a series of chimeric {dollar}\alpha{dollar}-factor receptors in which certain regions of Saccharomyces cerevisiae {dollar}\alpha{dollar}-factor receptor were replaced by corresponding regions of the Saccharomyces kluyveri {dollar}\alpha{dollar}-factor receptor. Two noncontiguous regions of the {dollar}\alpha{dollar}-factor receptor encompassing the first three and the last two transmembrane segments along with the associated loop regions were initially identified as controlling selectivity for S. cerevisiae vs. S. kluyveri {dollar}\alpha{dollar}-factor peptides. These regions confer selectivity to both ligand binding and activation of cellular responses.;Further characterization involved a more detailed mapping of selectivity determinants. Replacement of the juxtamembrane region near the N-terminal domain of the receptor had a marked influence on ligand binding specificity, but did not affect the specificity for activation. In contrast, a double substitution in the third extracellular loop of the receptor changed the activation specificity, leaving the ligand binding specificity unaltered. The rest of the regions identified as ligand specificity determinants are understood to control the selectivity for both ligand binding and activation of cellular responses.