G-BETA-GAMMA BINDS DIRECTLY TO THE G-PROTEIN-GATED K+ CHANNEL, I-KACH

The cardiac G protein-gated K+ channel, I-KACh, is activated by application of purified and recombinant beta and gamma subunits (G beta gamma) of heterotrimeric G proteins to excised inside-out patches from atrial membranes (Logothetis, D. E., Kurachi, Y., Galper, J., Neer, E., and Clapham, D. E., (1987) Nature 325, 321-326; Wickman, K., Iniguez-Lluhi, J., Davenport, P., Taussig, R. A., Krapivinsky, G. B., Linder, M. E., Gilman, A., and Clapham, D. E. (1994) Nature 368, 255-257). Cardiac I-KACh is composed of two inward rectifier K+ channel subunits, GIRK1 and CIR (Krapivinsky, G., Gordon, E., Wickman, K., Velimirovic, B., Krapivinsky, L., and Clapham, D. E. (1995) Nature 374, 135-141). We show that G beta gamma directly binds to immunoprecipitated cardiac I-KACh as well as to recombinant CIR and GIRK1 subunits, with dissociation constants (K-d) of 55, 50, and 125 nM, respectively. In each case, binding appeared specific as judged by competition of unlabeled G beta gamma with radiolabeled G beta gamma and inhibition of binding by antigenic peptide or G alpha-GDP, but not G alpha-GTP gamma S (guanosine 5'-3-O-(thio)triphosphate). In contrast, G alpha (GTP gamma S- or GDP-bound) did not bind to the native channel. We conclude that G beta gamma binds directly and specifically to I-KACh via interactions with both CIR and GIRK1 subunits to gate the channel.