Agonists induce conformational changes in transmembrane domains III and VI of the beta(2) adrenoceptor

Agonist binding to G protein-coupled receptors is believed to promote a conformational change that leads to the formation of the active receptor state, However, the character of this conformational change which provides the important link between agonist binding and G protein coupling is not known, Here we report evidence that agonist binding to the beta(2) adrenoceptor induces a conformational change around (125)Cys in transmembrane domain (TM) III and around (285)Cys in TM VI, A series of mutant beta(2) adrenoceptors with a limited number of cysteines available for chemical derivatization were purified, site-selectively labeled with the conformationally sensitive, cysteine-reactive fluorophore IANBD and analyzed by fluorescence spectroscopy, Like the wild-type receptor, mutant receptors containing (125)Cys and/or (285)Cys showed an agonist-induced decrease in fluorescence, while no agonist-induced response was observed in a receptor where these two cysteines were mutated, These data suggest that IANBD bound to (125)Cys and (285)Cys are exposed to a more polar environment upon agonist binding, and indicate that movements of transmembrane segments III and VI are involved in activation of G protein-coupled receptors.