G-protein alpha-subunit expression, myristoylation, and membrane association in COS cells.
Myristoylation of seven different alpha subunits of guanine nucleotide-binding regulatory proteins (G proteins) was examined by expressing these proteins in monkey kidney COS cells. Metabolic labeling studies of cells transfected with cytomegalovirus-based expression vectors indicated that [3H]myristate was incorporated into alpha i1, alpha i2, alpha i3, alpha 0, alpha t, and alpha z but not alpha s subunits. The role of myristoylation in the association of alpha subunits with membranes was analyzed by site-directed mutagenesis and by substitution of myristate with a less hydrophobic analog, 10-(propoxy)decanoate (11-oxamyristate). Myristoylation of alpha 0 was blocked when an alanine residue was substituted for its amino-terminal glycine, as was association of the protein with membranes. Substitution of the myristoyl group with 11-oxamyristate affected the cellular distribution of a subset of acylated alpha subunits. The results are consistent with a model wherein the hydrophobic interaction of myristate with the bilayer permits continued association of the protein with the plasma membrane when G-protein alpha subunits dissociate from beta gamma.
Mumby
S M
SM
Department of Pharmacology, University of Texas Southwestern Medical Center, Dallas 75235.
Heukeroth
R O
RO
Gordon
J I
JI
Gilman
A G
AG
D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism*; D003587:Cytomegalovirus / Q000235:genetics; D004247:DNA / Q000235:genetics; D019204:GTP-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D005822:Genetic Vectors; D007700:Kinetics; D046911:Macromolecular Substances; D009154:Mutation; D019814:Myristic Acid; D009227:Myristic Acids / Q000378:metabolism*; D010957:Plasmids; D011499:Protein Processing, Post-Translational; D014162:Transfection
D046911:Macromolecular Substances; D009227:Myristic Acids; D019814:Myristic Acid; D004247:DNA; D019204:GTP-Binding Proteins