Fasciola hepatica: heterologous expression and functional characterization of a thioredoxin peroxidase.
A Fasciola hepatica cDNA clone of 779 bp was isolated from an adult worm cDNA expression library by immunological screening using a rabbit serum against the excretory-secretory antigens. The nucleotide sequence of the cDNA revealed the presence of an open reading frame of 582 bp which encoded a 194-amino-acid-residue polypeptide (M(r) 21,723 Da) showing a high degree of homology to thioredoxin peroxidases. This putative antioxidant protein gene was expressed in Escherichia coli as a GST fusion protein. The recombinant fusion protein showed in vitro antioxidant properties and protected rabbit muscle enolase and E. coli glutamine synthetase from inactivation by nonenzymatic Fe(3+)/O(2)/DTT or Fe(3+)/O(2)/ascorbate metal-catalyzed oxidation systems.
Salazar-Calderón
M
M
Departamento de Bioquímica y Biología Molecular, Instituto Universitario de Biotecnología de Asturias (CSIC), Universidad de Oviedo, Oviedo, 33006, Spain.
Martín-Alonso
J M
JM
Ruiz de Eguino
A D
AD
Casais
R
R
Marín
M S
MS
Parra
F
F
D000595:Amino Acid Sequence; D000818:Animals; D000975:Antioxidants / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D001483:Base Sequence; D015152:Blotting, Northern; D015139:Blotting, Southern; D015153:Blotting, Western; D002417:Cattle; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000737:chemistry; D018509:DNA, Helminth / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D005210:Fasciola hepatica / Q000201:enzymology* / Q000235:genetics; D015870:Gene Expression; D008969:Molecular Sequence Data; D009363:Neoplasm Proteins; D016366:Open Reading Frames; D010544:Peroxidases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D054464:Peroxiredoxins; D012333:RNA, Messenger / Q000032:analysis; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid
D000975:Antioxidants; D018076:DNA, Complementary; D018509:DNA, Helminth; D009363:Neoplasm Proteins; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D010544:Peroxidases; D054464:Peroxiredoxins