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 GTPgammaS (guanosine 5'-O-(thio)triphosphate) at 1 muM free Mg2+, in the presence of phospholipids. Unmyristoylated ARF(GTPgammaS) 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 ARF(GDP) with its membrane-bound exchange factor rather than for the association of ARF(GTP) with lipid membranes. Phospholipid-bound ARF(GTPgammaS) pan also stably interact with and activate the catalytic subunit of cholera toxin, suggesting that ARF(GTP) provides a membrane anchor for cholera toxin and thereby facilitates its access to membrane-bound substrates.