Myristoylation is not required for GTP-dependent binding of ADP-ribosylation factor ARF1 to phospholipids.
Membrane binding of ADP-ribosylation factors (ARFs) is GTP-dependent and seems to require amino-terminal myristoylation. Recently it has been proposed that myristoylation is needed not for the activation of ARF by GTP but for its subsequent association to membranes. Here we show that unmyristoylated bovine ARF1, expressed in bacteria, can be efficiently loaded with GTP gamma S (guanosine 5'-O-(thio)triphosphate) at 1 microM free Mg2+, in the presence of phospholipids. Unmyristoylated ARFGTP gamma S cosediments with phospholipid vesicles and totally binds to phospholipid-cholate micelles, as seen by gel filtration chromatography. We therefore propose that, in vivo, myristoylation is required for the interaction of ARFGDP with its membrane-bound exchange factor rather than for the association of ARFGTP with lipid membranes. Phospholipid-bound ARFGTP gamma S can also stably interact with and activate the catalytic subunit of cholera toxin, suggesting that ARFGTP provides a membrane anchor for cholera toxin and thereby facilitates its access to membrane-bound substrates.
Franco
M
M
Centre National de la Recherche Scientifique, Institut de Pharmacologie moléculaire et cellulaire, Valbonne, France.
Chardin
P
P
Chabre
M
M
Paris
S
S
D020823:ADP-Ribosylation Factor 1; D020727:ADP-Ribosylation Factors; D000818:Animals; D001483:Base Sequence; D001692:Biological Transport; D002352:Carrier Proteins / Q000378:metabolism*; D002417:Cattle; D002462:Cell Membrane / Q000378:metabolism; D002772:Cholera Toxin / Q000378:metabolism; D018076:DNA, Complementary; D019204:GTP-Binding Proteins / Q000378:metabolism*; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000378:metabolism*; D008823:Micelles; D008969:Molecular Sequence Data; D019814:Myristic Acid; D009227:Myristic Acids / Q000378:metabolism*; D009838:Oligodeoxyribonucleotides; D010743:Phospholipids / Q000378:metabolism*
D002352:Carrier Proteins; D018076:DNA, Complementary; D008823:Micelles; D009227:Myristic Acids; D009838:Oligodeoxyribonucleotides; D010743:Phospholipids; D019814:Myristic Acid; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D006160:Guanosine Triphosphate; D002772:Cholera Toxin; D019204:GTP-Binding Proteins; D020823:ADP-Ribosylation Factor 1; D020727:ADP-Ribosylation Factors