RESTRICTING MOBILITY OF G(S)ALPHA RELATIVE TO THE BETA(2)-ADRENOCEPTOR ENHANCES ADENYLATE-CYCLASE ACTIVITY BY REDUCING G(S)ALPHA GTPASE ACTIVITY

The beta(2)-adrenoceptor (beta(2)AR) activates the G-protein G(s)alpha to stimulate adenylate cyclase (AC). Fusion of the beta(2)AR C-termin us to the N-terminus of G(s)alpha (producing beta(2)ARG(s)alpha) marke dly increases the efficiency of receptor/G-protein coupling compared w ith the non-fused state. This increase in coupling efficiency can be a ttributed to the physical proximity of receptor and G-protein. To dete rmine the optimal length for the tether between receptor and G-protein we constructed fusion proteins from which 26 [beta(2)AR(Delta 26)G(s) alpha] or 70 [beta(2)AR(Delta 70)G(s)alpha] residues of the beta(2)AR C-terminus had been deleted and compared the properties of these fusio n proteins with the previously described beta(2)ARG(s)alpha. Compared with beta(2)ARG(s)alpha, basal and agonist-stimulated GTP hydrolysis w as markedly decreased in beta(2)AR(Delta 70)G(s)alpha, whereas the eff ect of the deletion on binding of guanosine 5'-[gamma-thio]triphosphat e (GTP[S]) was relatively small. Surprisingly, deletions did not alter the efficiency of coupling of the beta(2)AR to G(s)alpha as assessed by GTP[S]-sensitive high-affinity agonist binding. Moreover, basal and ligand-regulated AC activities in membranes expressing beta(2)AR(Delt a 70)G(s)alpha and beta(2)AR(Delta 26)G(s)alpha were higher than in me mbranes expressing beta(2)ARG(s)alpha. These findings suggest that res tricting the mobility of G(s)alpha relative to the beta(2)AR results i n a decrease in G-protein inactivation by CTP hydrolysis and thereby e nhanced activation of AC.