Role of Isoprenoid Lipids on the Heterotrimeric G Protein γ Subunit in Determining Effector Activation*

Post-translational prenylation of heterotrimeric G protein γ subunits is essential for high affinity α-βγ and α-βγ-receptor interactions, suggesting that the prenyl group is an important domain in the βγ dimer. To determine the role of the prenyl modification in the interaction of βγ dimers with effectors, the CAAX (where A indicates alipathic amino acid) motifs in the γ₁, γ₂, and γ₁₁ subunits were altered to direct modification with different prenyl groups. Six recombinant βγ dimers were overexpressed in baculovirus-infected Sf9 insect cells, purified, and examined for their ability to stimulate three phospholipase C-β isozymes and type II adenylyl cyclase. The native β₁γ₂ dimer (γ subunit modified with geranylgeranyl) is more potent and effective in activating phospholipase C-β than either the β₁γ₁ (farnesyl) or the β₁γ₁₁ (farnesyl) dimers. However, farnesyl modification of the γ subunit in the β₁γ₂dimer (β₁γ_2-L71S) caused a decrement in its ability to activate phospholipase C-β. In contrast, both the β₁γ_1-S74L (geranylgeranyl) and the β₁γ_11-S73L (geranylgeranyl) dimers were more active than the native forms. The β₁γ₂dimer activates type II adenylyl cyclase about 12-fold; however, neither the β₁γ₁ nor the β₁γ₁₁ dimers activate the enzyme. As was the case with phospholipase C-β, the β₁γ_2-L71S dimer was less able to activate adenylyl cyclase than the native β₁γ₂dimer. Interestingly, neither the β₁γ_1-S74Lnor the β₁γ_11-S73L dimers stimulated adenylyl cyclase. The results suggest that both the amino acid sequence of the γ subunit and its prenyl group play a role in determining the activity of the βγ-effector complex.