Mutant G protein α subunit activated by Gβγ:: A model for receptor activation?

How receptors catalyze exchange of CTP for GDP bound to the G alpha subunit of trimeric G proteins is not known. One proposal is that the receptor uses the C protein's beta gamma heterodimer as a lever, tilting it to pull open the guanine nucleotide binding pocket of G alpha. To test this possibility, we designed a mutant G alpha that would bind to beta gamma in the tilted conformation. To do so, we excised a helical turn (four residues) from the N-terminal region of alpha (s), the a subunit of G(s), the stimulatory regulator of adenylyl cyclase. In the presence, but not in the absence, of transiently expressed beta (1) and gamma (2), this mutant (alpha (s)Delta), markedly stimulated cAMP accumulation. This effect depended on the ability of the coexpressed beta protein to interact normally with the lip of the nucleotide binding pocket of alpha (2)Delta. We substituted alanine for an aspartate in beta (1) that binds to a lysine (K206) in the lip of the a subunit's nucleotide binding pocket. Coexpressed with alpha (2)Delta and gamma (2), this mutant, beta1-D228A, elevated cAMP much less than did beta (1)-wild type; it did bind to alpha (2)Delta normally, however, as indicated by its unimpaired ability to target alpha (2)Delta to the plasma membrane. We conclude that beta gamma can activate alpha (s) and that this effect probably involves both a tilt of beta gamma relative to alpha (s) and interaction of beta with the lip of the nucleotide binding pocket. We speculate that receptors use a similar mechanism to activate trimeric G proteins.