Receptor−G Protein γ Specificity: γ11 Shows Unique Potency for A1Adenosine and 5-HT1AReceptors

Abstract

G protein coupled receptors activate signal transducing guanine nucleotide-binding proteins (G proteins), which consist of an α subunit and a βγ dimer. Whole cell studies have reported that receptors signal through specific βγ subtypes. Membrane reconstitution studies with the adenosine A₁ and α_2A adrenergic receptors have reached a similar conclusion. We aimed to test the generality of this finding by comparing the γ subtype specificity for four G_i-coupled receptors: α_2A adrenergic; A1 adenosine (A₁-R); 5-hydroxytryptamine_1A (5-HT_1A-R); mu opioid. Membranes were reconstituted with Gα_i1 and five γ subtypes (dimerized to β1). Using a sensitive α-βγ binding assay, we show that all recombinant βγ (except β1γ1) had comparable affinity for α_i1. Using high affinity agonist binding as a measure of receptor−G protein coupling, βγ-containing γ11 was the most potent for A₁-R and 5-HT_1A-R (p < 0.05, one way ANOVA) while γ7 was most potent for the other two receptors. γ11 was 3−8-fold more potent for the A₁-R than were the other γ subtypes. Also, γ11 was 2−8-fold more potent for A₁-R than at the other receptors, suggesting a unique coupling specificity of the A₁-R for γ11. In contrast, the discrimination by receptors for the other βγ subtypes (β1 and γ1, γ2, γ7, and γ10) was limited (2−3-fold). Thus the exquisite βγ specificity of individual receptors reported in whole cell studies may depend on in vivo mechanisms beyond direct receptor recognition of βγ subtypes.