Role of the chaperonin CCT/TRiC complex in G protein βγ-dimer assembly

G beta gamma dimer formation occurs early in the assembly of heterotrimeric G proteins. On nondenaturing (native) gels, in vitro translated, S-35-labeled G gamma subunits traveled primarily according to their pI and apparently were not associated with other proteins. In contrast, in vitro translated, S-35-labeled G beta subunits traveled at a high apparent molecular mass (similar to 700 kDa) and co-migrated with the chaperonin CCT complex ( also called TRiC). Different FLAG-G beta isoforms coprecipitated CCT/TRiC to a variable extent, and this correlated with the ability of the different G beta subunits to efficiently form dimers with G gamma. When translated G gamma was added to translated G gamma, a new band of low apparent molecular mass (similar to 50 kDa) was observed, which was labeled by either S-35-labeled G gamma or G gamma, indicating that it is a dimer. Formation of the G beta gamma dimer was ATP-dependent and inhibited by either adenosine 5'-O-(thiotriphosphate) or aluminum fluoride in the presence of Mg2+. This inhibition led to increased association of G gamma with CCT/TRiC. Although G gamma did not bind CCT/TRiC, addition of G gamma to previously synthesized G beta caused its release from the CCT/TRiC complex. We conclude that the chaperonin CCT/TRiC complex binds to and folds G beta subunits and that CCT/TRiC mediates G beta gamma dimer formation by an ATP-dependent reaction.