The G protein β5 subunit interacts selectively with the Gq α subunit

The diversity in the heterotrimeric G protein alpha, beta, and gamma subunits may allow selective protein-protein interactions and provide specificity for signaling pathways. We examined the ability of five a subunits (alpha(i1), alpha(i2), alpha(o), alpha(s), and alpha(q),) to associate with three beta subunits (beta(1), beta(2), and beta(5)) dimerized to a gamma(2) subunit containing an amino-terminal hexahistidine-FLAG affinity tag (gamma(2HF)). Sf9 insect cells were used to overexpress the recombinant proteins. The hexahistidine-FLAG sequence does not hinder the function of the beta(1) gamma(2HF) dimer as it can be specifically eluted from an alpha(i1)-agarose column with GDP and AlF4-, and purified beta(1) gamma(2HF) dimer stimulates type II adenylyl cyclase. The beta(1) gamma(2HF) and beta(2) gamma(2HF) dimers immobilized on an anti-FLAG affinity column bound all five alpha subunits tested, whereas the beta(5) gamma(2HF) dimer bound only alpha(q). The ability of other alpha subunits to compete with the alpha(q) subunit for binding to the beta(5) gamma(2HF) dimer was tested. Addition of increasing amounts of purified, recombinant alpha(i1) to the alpha(q) in a Sf9 cell extract did not decrease the amount of alpha(q) bound to the beta(5) gamma(2HF) column. When G proteins in an extract of brain membranes were activated with GDP and AlF4- and deactivated in the presence of equal amounts of the beta(1) gamma(2HF) or beta(5) gamma(2HF) dimers, only alpha(q) bound to the beta(5) gamma(2HF) dimer. The alpha(q) beta(5) gamma(2HF) interaction on the column was functional as GDP, and AlF4- specifically eluted cu, from the column. These results indicate that although the beta(1) and beta(2) subunits interact with alpha subunits from the alpha(i), alpha(s), and alpha(q) families, the structurally divergent beta(5) subunit only interacts with alpha(q).