Molecular cloning and characterization of a 2-Cys peroxiredoxin from Taenia solium.
A Taenia solium 2-Cys peroxiredoxin (Ts2-CysPrx) clone was isolated from a T. solium adult cDNA library. The clone encodes a polypeptide comprising 197 amino acids with a predictive Mr = 21,836. It has the 2 classical cysteine domains from the typical 2-Cys peroxiredoxins, and its primary amino acid sequence shows higher identity with 2 Echinococcus 2-Cys peroxiredoxins. Northern and Southern blot hybridizations exhibit an mRNA with a size of -1.0 kb, encoded by 1 gene. Ts2-CysPrx was expressed in Escherichia coli and purified by anion-exchange chromatography. Biochemical analysis showed Ts2-CysPrx is a dimer composed by monomers of -22 kDa that presented activity with hydrogen peroxide (H2O2) and cumene hydroperoxide. It presented the catalytic mechanism for a typical 2-CysPrx because the homodimeric oxidized form is reduced to a monomeric form by thioredoxin (Trx) and by dithiothreitol (DTT) and was converted to a homodimeric oxidized form by H2O2. Western blot studies using antibodies against Ts2-CysPrx revealed that the protein is expressed during the entire T. solium life cycle, as in other Taenia species. Immunohistochemical studies indicated that Ts2-CysPrx is localized on the tegument and in tegumentary and muscle cells of cysticerci. We also show that T. crassiceps cysticerci can tolerate H2O2 levels of 2.5 mM for 2.5 hr.
Molina-López
José
J
Departamento de Microbiología y Parasitología, Facultad de Medicina, Universidad Nacional Autónoma de Mexico. Ciudad Universitaria, México DF 04510, México.
Jiménez
Lucía
L
Ochoa-Sánchez
Alicia
A
Landa
Abraham
A
D000595:Amino Acid Sequence; D000818:Animals; D000909:Antibodies, Helminth / Q000276:immunology; D001483:Base Sequence; D015152:Blotting, Northern; D015139:Blotting, Southern; D015153:Blotting, Western; D003001:Cloning, Molecular; D003545:Cysteine / Q000737:chemistry / Q000378:metabolism; D003552:Cysticercus / Q000201:enzymology / Q000235:genetics; D018076:DNA, Complementary / Q000737:chemistry / Q000302:isolation & purification; D018509:DNA, Helminth / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D015723:Gene Library; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism / Q000494:pharmacology; D051379:Mice; D008969:Molecular Sequence Data; D010544:Peroxidases / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D054464:Peroxiredoxins; D012333:RNA, Messenger / Q000032:analysis; D011817:Rabbits; D016415:Sequence Alignment; D013552:Swine; D041201:Taenia solium / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics
D000909:Antibodies, Helminth; D018076:DNA, Complementary; D018509:DNA, Helminth; D012333:RNA, Messenger; D006861:Hydrogen Peroxide; D010544:Peroxidases; D054464:Peroxiredoxins; D003545:Cysteine