Extensive purification from Acanthamoeba castellanii of a microtubule-dependent translocator with microtubule-activated Mg2+-ATPase activity.
A protein which supported MgATP-dependent movement of latex beads from the minus to the plus end of microtubules and which had microtubule-activated Mg2+-ATPase was purified from Acanthamoeba castellanii. At concentrations as low as 0.6 micrograms ml-1, the translocator supported movement of beads at a rate of 3 to 4 micron s-1. The translocator protein had a Ca2+-ATPase activity of 1.7 mumol min-1 mg-1 and a Mg2+-ATPase activity of about 0.03 mumol min-1 mg-1 in the absence of microtubules. The Mg2+-ATPase in the presence of microtubules had a Vmax of 3.4 mumol min-1 mg-1; half-maximal Mg2+-ATPase activity required only 0.45 microM microtubules (concentration of dimer subunits). The highly purified native protein had a Stokes radius of 8.5 nm, and three polypeptides of Mr 134,000, 139,000, and 147,000 were associated with the fractions that had maximum translocator and ATPase activities.
D000048:Acanthamoeba / Q000201:enzymology*; D000818:Animals; D017301:Ca(2+) Mg(2+)-ATPase / Q000302:isolation & purification* / Q000378:metabolism; D007447:Invertebrate Hormones / Q000302:isolation & purification* / Q000378:metabolism; D007700:Kinetics; D008868:Microtubule Proteins / Q000302:isolation & purification* / Q000378:metabolism; D008870:Microtubules / Q000201:enzymology* / Q000648:ultrastructure
D007447:Invertebrate Hormones; D008868:Microtubule Proteins; D017301:Ca(2+) Mg(2+)-ATPase