Hormonal inhibition of adenylate cyclase is mediated by inhibitory receptors and a guanyl nucleotide-binding coupling protein, termed Gi. Similarly, transducin (T), a guanyl nucleotide-binding protein, mediates activation of cGMP phosphodiesterase by the retinal photon receptor, rhodopsin. Gi and T are both heterotrimers consisting of alpha, beta, and gamma subunits; Gi alpha and G beta are similar to T alpha and T beta, respectively. T alpha hydrolyzes GTP in the presence of photolyzed, but not dark, rhodopsin and T beta gamma. Gi alpha and G beta gamma substituted for T alpha and T beta gamma to yield active hybrid complexes, T alpha G beta gamma and Gi alpha T beta gamma. In the absence of T components, rhodopsin-dependent GTPase activity of Gi alpha G beta gamma was observed. Pertussis toxin ADP-ribosylates both T alpha and Gi alpha; ADP-ribosylation of Gi alpha was negligible in the absence of G beta gamma. With G beta gamma, photolyzed, but not dark, rhodopsin unhibited ADP-ribosylation of Gi alpha. In the presence of G beta gamma and photolyzed rhodopsin, GDP and GDP beta S, but not Gpp(NH)p and GTP gamma S, increased the ADP-ribosylation of Gi alpha. The requirements for ADP-ribosylation of Gi alpha by pertussis toxin were similar to those for ADP-ribosylation of T alpha. Rhodopsin appears to interact with Gi in a manner similar to the inhibitory hormone receptors; photolyzed rhodopsin, the active species, corresponds to the agonist-occupied receptor, while dark rhodopsin, the inactive species, can be equated to the free receptor.(ABSTRACT TRUNCATED AT 250 WORDS)