Partial purification and kinetic properties of cytoplasmic malate dehydrogenase from ovine liver Echinococcus granulosus protoscolices.
Cytoplasmic malate dehydrogenase from ovine liver Echinococcus granulosus protoscolices was purified 22-fold by QAE- and SP-Sephadex chromatography. The pH optimum of the enzyme was 8.0 in either Tris-HCI or barbital buffer. The Km values of oxaloacetate and NADH were 0.400 +/- 0.018 and 0.410 +/- 0.038 mM, respectively. The enzyme lost about 90% of its activity when heated for 2 min at 65 degrees C. A 61.4% inhibition of the enzyme was noted at 4 mM concentration of diethyl pyrocarbonate. A 3 mM concentration of fructose 1,6-diphosphate inhibited the enzyme by 76.5%. The inhibition was non-competitive with respect to NADH with a Ki value of 0.85 mM. A 75% inhibition of the enzyme was noted at 1 mM concentration of mebendazole that inhibited the enzyme upon competing with NADH with a Ki value of 0.176 mM. A 2-mM concentration of citrate almost doubled the enzyme activity. The enzyme was inhibited at high concentrations of either substrate. The enzyme was not inhibited by p-hydroxymercuribenzoate or fumarate. The enzyme was absolutely specific for NADH as a cofactor. The properties of this enzyme are compared with those of the enzyme from the host liver, the cyst fluid and some other animal sources. The results are discussed in terms of the differences among the properties of the host liver, the cyst fluid and the protoscolices enzymes. The biochemical basis for the use of mebendazole in the treatment of echinococcosis is also elucidated.
Vessal
M
M
Department of Biochemistry, Shiraz University of Medical Sciences, Iran.
Tabei
S M
SM
D000818:Animals; D000969:Antinematodal Agents / Q000494:pharmacology; D001826:Body Fluids / Q000201:enzymology; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D003593:Cytoplasm / Q000201:enzymology*; D004443:Echinococcosis / Q000188:drug therapy; D004446:Echinococcus / Q000201:enzymology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D007700:Kinetics; D008099:Liver / Q000469:parasitology*; D008291:Malate Dehydrogenase / Q000037:antagonists & inhibitors / Q000302:isolation & purification* / Q000378:metabolism; D008463:Mebendazole / Q000494:pharmacology; D012756:Sheep / Q000469:parasitology*
D000969:Antinematodal Agents; D004791:Enzyme Inhibitors; D008463:Mebendazole; D008291:Malate Dehydrogenase