Role of isoprenoid lipids on the heterotrimeric G protein γ subunit in determining effector activation

Post-translational prenylation of heterotrimeric G protein gamma subunits is essential for high affinity alpha-beta gamma and alpha-beta gamma-receptor interactions, suggesting that the prenyl group is an important domain in the beta gamma dimer. To determine the role of the prenyl modification in the interaction of beta gamma dimers with effecters, the CAAX (where A indicates alipathic amino acid) motifs in the gamma(1), gamma(2), and y(11) subunits were altered to direct modification with different prenyl groups. Six recombinant beta gamma dimers were overexpressed in baculovirus-infected Sf9 insect cells, purified, and examined for their ability to stimulate three phospholipase C-beta isozymes and type II adenylyl cyclase, The native beta(1)gamma(2) dimer (gamma subunit modified with geranylgeranyl) is more potent and effective in activating phospholipase C-beta than either the beta(1)gamma(1) (farnesyl) or the beta(1)gamma(11) (farnesyl) dimers. However, farnesyl modification of the gamma subunit in the beta(1)gamma(2) dimer (beta(1)gamma(2-L71S)) caused a decrement in its ability to activate phospholipase C-beta. In contrast, both the beta 1 gamma(1-S74L) (geranylgeranyl) and the beta(1)gamma(11-S73L) (geranylgeranyl) dimers were more active than the native forms. The beta(1)gamma(2) dimer activates type II adenylyl cyclase about 12-fold; however, neither the beta(1)gamma(1) nor the beta(1)gamma(11) dimers activate the enzyme. As was the case with phospholipase C-P, the beta(1)gamma(2-L71S) dimer was less able to activate adenylyl cyclase than the native beta(1)gamma(2) dimer. Interestingly, neither the beta(1)gamma(1-S74L) nor the beta(1)gamma(11-S73L) dimers stimulated adenylyl cyclase. The results suggest that both the amino acid sequence of the gamma subunit and its prenyl group play a role in determining the activity of the beta gamma-effector complex.