Purification and Characterization of Recombinant G16αFrom Sf9 Cells: Activation of Purified Phospholipase C Isozymes by G-Protein α Subunits

A cDNA encoding G16α, the α subunit of a heterotrimeric guanine nucleotide-binding protein, was expressed in Sf9 cells using recombinant baculovirus. G16αin membrane extracts of Sf9 cells activated phospholipase C-β1 (PLC-β1) in the presence of guanosine 5'-[γ-thio]triphosphate; the system could not be activated by Al3+, Mg2+, and F-. The G16αin the cytosolic fraction of Sf9 cells did not stimulate PLC-β1. Concurrent expression of the G-protein βγ subunit complex increased the amount of G16αin Sf9 cell membranes. The guanosine 5'-[γ-thio]triphosphate-activated form of G16αwas purified from cholate extracts of membranes from cells expressing G16α, and the G-protein β2and γ2subunits. G16αactivated PLC-β1, PLC-β2, and PLC-β3 in a manner essentially indistinguishable from that of Gqα. G16α-mediated activation of PLC-β1 and PLC-β3 greatly exceeded that of PLC-β2. G16αdid not activate PLC-γ1 or PLC-δ1. Thus, two distantly related members of the Gqαfamily, Gqαand G16α, have the same ability to activate the known isoforms of PLC-β.