Effect of myristoylation on GTP-dependent binding of ADP-ribosylation factor to Golgi.
ADP-ribosylation factors (ARFs), a family of approximately 20-kDa guanine nucleotide-binding proteins that activate cholera toxin ADP-ribosyltransferase in vitro, have been implicated in intracellular protein trafficking and are thought to cycle between cytosolic and membrane compartments. Although isolated predominantly as soluble proteins, ARFs associate with membranes and phospholipids in a GTP-dependent manner. In contrast to other small GTP-binding proteins, ARFs are NH2 terminally myristoylated. Using a bacterial expression system, recombinant myristoylated and non-myristoylated human ARF5 were produced to investigate the role of myristoylation in its association with Golgi. The recombinant ARFs (myristoylated and non-myristoylated) exhibited similar biochemical activity as measured by GTP binding and in vitro activation of cholera toxin. Myristoylated ARF5, however, demonstrated a temperature- and GTP-dependent association with Golgi membranes, whereas non-myristoylated ARF did not bind to Golgi under any of the experimental conditions. These data indicate that myristoylation is necessary, although not sufficient, for membrane attachment, but is not necessary for activation of cholera toxin.
Haun
R S
RS
Laboratory of Cellular Metabolism, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland 20892.
Tsai
S C
SC
Adamik
R
R
Moss
J
J
Vaughan
M
M
D020727:ADP-Ribosylation Factors; D001483:Base Sequence; D002352:Carrier Proteins / Q000378:metabolism*; D004271:DNA, Fungal; D019204:GTP-Binding Proteins / Q000378:metabolism*; D006056:Golgi Apparatus / Q000378:metabolism*; D006160:Guanosine Triphosphate / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D009227:Myristic Acids / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D013696:Temperature
D002352:Carrier Proteins; D004271:DNA, Fungal; D009227:Myristic Acids; D006160:Guanosine Triphosphate; D019204:GTP-Binding Proteins; D020727:ADP-Ribosylation Factors