A cysteine-11 to serine mutant of G alpha12 impairs activation through the thrombin receptor.

We have recently reported that G alpha12 is acylated with palmitic acid [Veit et al., FEBS Lett. 339 (1994) 160-164]. Here we identify cysteine 11 as the sole palmitoylation site and assess the function of G alpha12 palmitoylation after expression of wild type and acylation-deficient mutant in insect cells. Our experimental approach yielded the following results. (1) Palmitoylation of G alpha12 has no influence on the subunit interactions. (2) Palmitoylation promotes membrane binding of G alpha12 when this protein is expressed alone. Membrane attachment of the heterotrimer occurs independent of the presence of fatty acids in G alpha12. (3) Assays for agonist-stimulated binding of [35S]GTPgammaS after expression of the human thrombin receptor (PAR1) along with G alpha12 and the betagamma subunits revealed a 70% inhibition with the palmitoyl-deficient mutant.