The G protein beta5 subunit interacts selectively with the Gq alpha 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 alpha subunits (alphai1, alphai2, alphao, alphas, and alphaq) to associate with three beta subunits (beta1, beta2, and beta5) dimerized to a gamma2 subunit containing an amino-terminal hexahistidine-FLAG affinity tag (gamma2HF). Sf9 insect cells were used to overexpress the recombinant proteins. The hexahistidine-FLAG sequence does not hinder the function of the beta1gamma2HF dimer as it can be specifically eluted from an alphai1-agarose column with GDP and AlF4-, and purified beta1gamma2HF dimer stimulates type II adenylyl cyclase. The beta1gamma2HF and beta2gamma2HF dimers immobilized on an anti-FLAG affinity column bound all five alpha subunits tested, whereas the beta5gamma2HF dimer bound only alphaq. The ability of other alpha subunits to compete with the alphaq subunit for binding to the beta5gamma2HF dimer was tested. Addition of increasing amounts of purified, recombinant alphai1 to the alphaq in a Sf9 cell extract did not decrease the amount of alphaq bound to the beta5gamma2HF 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 beta1gamma2HF or beta5gamma2HF dimers, only alphaq bound to the beta5gamma2HF dimer. The alphaq-beta5gamma2HF interaction on the column was functional as GDP, and AlF4- specifically eluted alphaq from the column. These results indicate that although the beta1 and beta2 subunits interact with alpha subunits from the alphai, alphas, and alphaq families, the structurally divergent beta5 subunit only interacts with alphaq.
Fletcher
J E
JE
Department of Pharmacology, Health Sciences Center, University of Virginia, Charlottesville, Virginia 22908, USA.
Lindorfer
M A
MA
DeFilippo
J M
JM
Yasuda
H
H
Guilmard
M
M
Garrison
J C
JC
D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D001921:Brain / Q000378:metabolism; D002417:Cattle; D002460:Cell Line; D019204:GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D018411:Spodoptera
D011994:Recombinant Proteins; D019204:GTP-Binding Proteins