A high affinity Ca2(+)-ATPase on the surface membrane of Leishmania donovani promastigote.
A Ca2(+)-dependent ATP-hydrolytic activity was detected in the crude membrane ghost of the promastigote or vector form of the protozoal parasite Leishmania donovani, the pathogen responsible for kala azar. The Ca2(+)-ATPase was purified to apparent homogeneity after solubilization with deoxycholate. The enzyme consists of two subunits of Mr = 51,000 and 57,000 and has an apparent molecular weight of 215,000 +/- 12,000. The enzyme activity is exclusively dependent on Ca2+, and the pure enzyme can hydrolyze 1.6 mumol of ATP/min/mg of protein. The apparent Km for Ca2+ is 35 nM, which is further reduced to 12 nM in the presence of heterologous calmodulin. The enzyme is sensitive to vanadate, but is insensitive to oligomycin and ouabain. The enzyme is strongly associated with the plasma membrane and has its catalytic site oriented toward the cytoplasmic face. The enzyme spans across the plasma membrane as surface labeling with radioiodine shows considerable radioactivity in the completely purified enzyme. The localization and orientation of this high affinity, calmodulin-sensitive Ca2(+)-ATPase suggest some role of this enzyme in Ca2+ movement in the life cycle of this protozoal parasite.
Ghosh
J
J
Enzyme Division, Indian Institute of Chemical Biology, Calcutta.
Ray
M
M
Sarkar
S
S
Bhaduri
A
A
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D001665:Binding Sites; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism*; D002147:Calmodulin / Q000494:pharmacology; D002384:Catalysis; D002462:Cell Membrane / Q000201:enzymology; D003593:Cytoplasm / Q000201:enzymology; D007457:Iodine Radioisotopes; D007700:Kinetics; D007893:Leishmania donovani / Q000201:enzymology* / Q000254:growth & development; D046911:Macromolecular Substances; D008970:Molecular Weight; D014638:Vanadates / Q000494:pharmacology
D002147:Calmodulin; D007457:Iodine Radioisotopes; D046911:Macromolecular Substances; D014638:Vanadates; D000255:Adenosine Triphosphate; D000252:Calcium-Transporting ATPases