Receptor selectivity between the G proteins Gα(12) and Gα(13) is defined by a single leucine-to-isoleucine variation

Despite recent advances in structural definition of GPCR–G protein complexes, the basis of receptor selectivity between G proteins remains unclear. The Gα(12) and Gα(13) subtypes together form the least studied group of heterotrimeric G proteins. G protein–coupled receptor 35 (GPR35) has been suggested to couple efficiently to Gα(13) but weakly to Gα(12). Using combinations of cells genome-edited to not express G proteins and bioluminescence resonance energy transfer–based sensors, we confirmed marked selectivity of GPR35 for Gα(13). Incorporating Gα(12)/Gα(13) chimeras and individual residue swap mutations into these sensors defined that selectivity between Gα(13) and Gα(12) was imbued largely by a single leucine-to-isoleucine variation at position G.H5.23. Indeed, leucine could not be substituted by other amino acids in Gα(13) without almost complete loss of GPR35 coupling. The critical importance of leucine at G.H5.23 for GPR35–G protein interaction was further demonstrated by introduction of this leucine into Gα(q), resulting in the gain of coupling to GPR35. These studies demonstrate that Gα(13) is markedly the most effective G protein for interaction with GPR35 and that selection between Gα(13) and Gα(12) is dictated largely by a single conservative amino acid variation.—Mackenzie, A. E., Quon, T., Lin, L.-C., Hauser, A. S., Jenkins, L., Inoue, A., Tobin, A. B., Gloriam, D. E., Hudson, B. D., Milligan, G. Receptor selectivity between the G proteins Gα(12) and Gα(13) is defined by a single leucine-to-isoleucine variation.