Specificity of receptor-G protein interactions: Discrimination of G_i subtypes by the D₂ dopamine receptor in a reconstituted system

The selectivity of D₂ dopamine receptor-guanine nucleotide-binding protein (G protein) coupling was studied by reconstitution techniques utilizing purified D₂ dopamine receptors from bovine anterior pituitary and resolved G proteins from bovine brain, bovine pituitary, and human erythrocyte. Titration of a fixed receptor concentration with varying G protein concentrations revealed two aspects of receptor-G protein coupling. First, G_i2 appeared to couple selectively with the D₂ receptor with ∼10-fold higher affinity than any other tested G_i subtype. Second, the G proteins differed in the maximal receptor-mediated agonist stimulation of the intrinsic GTPase activity. G_i2 appeared to be maximally stimulated by agonist-receptor complex with turnover numbers of ∼2 min^-1. The other G_i subtypes, G_i1 and G_i3, could be only partially activated, resulting in maximal rates of GTPase of ∼1 min^-1. Agonist-stimulated GTPase activity was not detected in preparations containing G_o from bovine brain. The differences in maximal agonist-stimulated GTPase rates observed among the G_i subtypes could be explained by differences in agonist-promoted guanyl nucleotide exchange. Both guanosine 5′-3-O-(thio)triphosphate (GTPγS) binding and GDP release parameters were enhanced 2-fold for the G_i2 subtype over the other G_i subtypes. These results suggest that even though several types of pertussis toxin substrate may exist in most tissues, a receptor may interact discretely with G proteins, thereby dictating signal transduction mechanisms.