Myristoylation of an inhibitory GTP-binding protein alpha subunit is essential for its membrane attachment.

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Abstract
We transfected COS cells with cDNAs for the .alpha. subunits of stimulatory and inhibitory GTP-binding proteins, .alpha.s and .alpha.i1, respectively, and immunoprecipitated the metabolically labeled products with specific peptide antibodies. Cells were separated into particulate and soluble fractions before immunoprecipitation; [35S]methionine-labeled .alpha.s and .alpha.i were both found primarily in the particulate fraction. [3H]Myristate was incorporated into endogenous and transfected .alpha.i but could not be detected in .alpha.s even when it was overexpressed. We converted the second residue, glycine, of .alpha.i1 into alanine by site-direction mutagenesis. Upon transfection of the mutant .alpha.i1 into COS cells, the [35S]methionine-labeled product was localized primarily to the soluble fraction, and, also unlike normal .alpha.i1, the mutant failed to incorporate [3H]myristate. The unmyristoylated mutant .alpha.i1 could still interact with the .beta.-.gamma. complex, since purified .beta..gamma. subunits promoted pertussis toxin-catalyzed ADP-ribosylation of both the normal and mutant .alpha.i1 subunits. These results indicate that myristoylation is critical for membrane attachment of .alpha.i but not .alpha.s subunits.