Characterization of transducin from bovine retinal rod outer segments. I. Separation and reconstitution of the subunits.
Transducin, a guanine nucleotide regulatory protein found in the bovine retinal rod outer segment, mediates the signal coupling between rhodopsin and a cyclic GMP phosphodiesterase. Previous studies have demonstrated that photolyzed rhodopsin catalyzed the exchange of GTP for GDP bound to transducin. The transducin-GTP complex, in turn, activates the phosphodiesterase. Purified transducin (T) has been resolved by omega-amino octylagarose chromatography into two functional subunits: T alpha (Mr approximately 39,000) and T beta gamma (Mr approximately 36,000 and approximately 10,000). The guanine nucleotide binding site is on the T alpha subunit. Neither the T alpha nor the T beta gamma subunit showed significant GTPase activity, Gpp(NH)p-GDP exchange, and ability to bind to rhodopsin when assayed in the presence of reconstituted membranes containing photolyzed rhodopsin. However, all the above activities were restored if the two subunits were recombined. Analysis of the reconstituted GTPase activity as a function of the T alpha subunit concentration revealed a linear relationship. On the other hand, GTPase activity rapidly saturated at T beta gamma concentration much lower than the T alpha concentration, indicating that the two subunits were dissociated during GTP hydrolysis. These findings strongly suggest that the T beta gamma subunit is an activator of the GTPase activity. Its mode of action is to enable the T alpha subunit to interact with rhodopsin effectively.
D000818:Animals; D002417:Cattle; D020558:GTP Phosphohydrolases / Q000378:metabolism; D006165:Guanylyl Imidodiphosphate / Q000378:metabolism; D007096:Imidoesters / Q000378:metabolism; D046911:Macromolecular Substances; D008565:Membrane Proteins / Q000032:analysis*; D008970:Molecular Weight; D010786:Photoreceptor Cells / Q000032:analysis*; D012374:Rod Cell Outer Segment / Q000032:analysis*; D015293:Transducin
D007096:Imidoesters; D046911:Macromolecular Substances; D008565:Membrane Proteins; C014539:ethyl acetimidate; D006165:Guanylyl Imidodiphosphate; D020558:GTP Phosphohydrolases; D015293:Transducin