Palmitoylation of a G protein α_i subunit requires membrane localization not myristoylation

Palmitoylation is a dynamic, post-translational modification of the amino terminus of heterotrimeric G protein α subunits. Since myristoylation, βγ interactions, and membrane attachment also involve the amino terminus of the G protein α_il subunit, we studied the relationships between palmitoylation and these events. Using COS cell transfection, the turnover of palmitate was slower on α_il subunits co-expressed with β and γ subunits than on the α_il subunit expressed alone. Mutation of cysteine 3 of α_il prevented [³H]palmitate but not [³H]myristate incorporation and decreased the membrane localization of this protein. This nonpalmitoylated mutant could form a heterotrimer with co-expressed βγ subunits which restored its membrane localization. A nonmyristoylated α_il mutant (glycine 2 to alanine) could incorporate [³H]palmitate when co-expressed with βγ subunits and localized to the membrane. The [³H]palmitate turnover of this nonmyristoylated mutant was more rapid than seen with the wild-type α_il subunit. While myristoylation is not required for palmitoylation, both myristoylation and βγ association can slow the turnover of palmitate on α_il. These results suggest that palmitoylation maintains the membrane attachment of the free α subunit and changes in βγ association could modulate palmitoylation during signaling.