Solubilization and kinetic characterization of mitochondrial adenosine triphosphatase from Leishmania donovani promastigotes.

Abstract Oligomycin-sensitive particulate ATPase (MB ATPase) from L. donovani promastigotes was solubilized by chloroform treatment. Polyacrylamide gel electrophoresis revealed several protein bands, with the major one possessing ATPase activity. The solubilized enzyme had Mg 2+ -ATPase and Ca 2− -ATPase but no K + -dependent alkaline phosphatase activity. The Mg 2+ -ATPase activity was stimulated by monovalent cations and was not sensitive to oligomycin. Hence it is referred to as F 1 ATPase. It had optimum activity at pH 7.6 and 30°C. The Arrhenius plot for MB ATPase was biphasic with activation energies ( E a ) of 16.2 and 3.4 kcal mol −1 , while F 1 ATPase exhibited a linear plot with E a = 10.1 kcal mol −1 . Lineweaver-Burk plots were biphasic with K m values of 0.17 and 1.25 mM for MB ATPase and 0.18 and 1.33 mM for F 1 ATPase. The enzyme could be preserved at −15°C in Tris-SO 4 2− -EDTA-ATP-glycerol ( t 1 2 = 20 days ).