G protein βγ subunits: Simplified purification and properties of novel isoforms

The β and γ subunits of heterotrimeric guanine nucleotide-binding regulatory proteins (G proteins) form tightly associated complexes. To examine functional differences among the large number of possible combinations of unique β and γ subunits, we have synthesized and characterized βγ complexes containing γ₅ and γ₇, two widely distributed γ subunits. When either γ₅ or γ₇ is expressed concurrently with β₁ or β₂ subunits in a baculovirus/Sf9 cell system, all four subunit complexes support pertussis toxin-catalyzed ADP-ribosylation of rG_iα1 (where "r" indicates recombinant), indicating formation of functional complexes. Each of the complexes was purified by subunit exchange chromatography, using the G203A mutant of rG_iα1 as the immobilized ligand. The purified preparations were compared with other recombinant βγ subunits, including β₁γ₁ and β₁γ₁, for their ability to modulate type I and II adenylyl cyclase activities; stimulate phosphoinositide-specific phospholipase Cβ; support pertussis toxin-catalyzed ADP-ribosylation of rG_iα1 and G_oα; and inhibit steady-state GTP hydrolysis catalyzed by G_sα, G_oα, and myristoylated rG_iα2. The results emphasize the unique properties of β₁γ₁. The properties of the complexes containing γ₅ or γ₇ were similar to each other and to those Of β₁γ₁.