Cloning, production and characterisation of a recombinant Cu/Zn superoxide dismutase from Taenia solium.
A full-length complementary DNA clone encoding a cytosolic Cu/Zn superoxide dismutase with a M(r) of 15,588 Da was isolated from a Taenia solium larvae complementary DNA library. Comparison analysis of its deduced amino acid sequence revealed a 71% identity with Schistosoma mansoni, 57.2-59.8% with mammalian and less than 54% with other helminth cytosolic Cu/Zn superoxide dismutase. The characteristic motifs and the amino acid residues involved in coordinating copper and zinc enzymatic function are conserved. The T. solium Cu/Zn superoxide dismutase was expressed in the pRSET vector. Enzymatic and filtration chromatographic analysis showed a recombinant enzyme with an activity of 2,941 U/mg protein and a native M(r) of 37 kDa. Inhibition assays using KCN, H(2)O(2), NaN(3) and SDS indicated that Cu/Zn is the metallic cofactor in the enzyme. Thiabendazole (500 microM) and albendazole (300 microM) completely inhibited the activity of T. solium Cu/Zn superoxide dismutase. Thiabendazole had no effect on bovine Cu/Zn superoxide dismutase; in contrast, albendazole had a moderate effect on it at same concentrations. Antibodies against T. solium Cu/Zn superoxide dismutase did not affect the enzymatic function; nevertheless, it cross reacts with several Taenia species, but not with trematodes, nematodes, pig, human and bovine Cu/Zn superoxide dismutase enzymes. Western blot analysis indicated the enzyme was expressed in all stages. These results indicate that T. solium possesses a Cu/Zn superoxide dismutase enzyme that can protect him from oxidant-damage caused by the superoxide anion.
Departamento de Microbiología y Parasitología, Facultad de Medicina, Edificio A, 2do piso, Universidad Nacional Autónoma de México, Ciudad Universitaria, Mexico D.F. 04510, Mexico.
Alejandro
A
Castellanos-González
Lucía
L
Jiménez
Abraham
A
Landa
D015766:Albendazole / Q000494:pharmacology; D000595:Amino Acid Sequence; D000818:Animals; D000871:Anthelmintics / Q000494:pharmacology; D015153:Blotting, Western; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011817:Rabbits; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013482:Superoxide Dismutase / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D041201:Taenia solium / Q000201:enzymology* / Q000235:genetics*; D013827:Thiabendazole / Q000494:pharmacology
D000871:Anthelmintics; D011994:Recombinant Proteins; D013482:Superoxide Dismutase; D015766:Albendazole; D013827:Thiabendazole