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Parasitology Research

, Volume 86, Issue 3, pp 200–206 | Cite as

Removal of hydrogen peroxide by a 1-cysteine peroxiredoxin enzyme of the filarial parasite Dirofilaria immitis

  • R. Chandrashekar
  • N. Tsuji
  • T. H. Morales
  • A. B. Carmody
  • V. O. Ozols
  • J. Welton
  • L. Tang
ORIGINAL PAPER

Abstract

Prior studies have shown that filarial nematodes can effectively metabolize hydrogen peroxide in excess of that generated by activated host cells. However, the mechanisms of H2O2 removal by the filarial parasites are unclear. Herein we report the results of studies carried out on the biochemical activity and on immunolocalization of a recombinant peroxiredoxin (Prx) enzyme from the dog filarial parasite Dirofilaria immitis. A full-length cDNA encoding a 1-Cys Prx enzyme from the dog heartworm D. immitis was expressed in Escherichia coli as a recombinant polyhistidine fusion protein (rDiPrx-1). rDiPrx-1 was capable of reducing H2O2 in the presence of dithiothreitol. The apparent kinetic constants determined for DiPrx-1 using H2O2 as a substrate were a Michaelis constant (Km) of 16.28 mM and a maximal velocity (vmax) of 16 μmol min−1. Consistent with the enzyme activity, D. immitis adult worms could detoxify exogenously added H2O2 in vitro. Antibodies to rDiPrx-1 identified a 27-kDa native antigen in parasite extracts and larval and adult excretory-secretory products. The antibodies were used to localize the native antigen to the lateral hypodermal chords of both male and female worms by immunohistochemistry. In addition, labeling was seen in the afibrillar muscle cells in male worms and in some areas of the uterine wall in female worms. Thus, DiPrx-1 is the first parasite Prx to be shown to detoxify exogenously added H2O2 in an in vitro system.

AbbreviationsL3 Third-stage larvae L4 Fourth-stage larvae SOD Superoxide dismutase GPx Glutathione peroxidase TSA Thiol-specific antioxidant Prx Peroxiredoxin TPx Thioredoxin peroxidase E-S products Excretory-secretory products MCO: Metal-ion-catalyzed oxidation rDiPrx-1 Recombinant D. immitis 1-Cys Pxr DTT Dithiothreitol 

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Copyright information

© Springer-Verlag Berlin Heidelberg 2000

Authors and Affiliations

  • R. Chandrashekar
    • 1
  • N. Tsuji
    • 1
  • T. H. Morales
    • 1
  • A. B. Carmody
    • 1
  • V. O. Ozols
    • 1
  • J. Welton
    • 1
  • L. Tang
    • 1
  1. 1.Heska Corporation, 1613 Prospect Parkway, Fort Collins, CO 80525, USA e-mail: Chandrr@Heska.com Tel.: +1-970-4937272; Fax: +1-970-493-7333US

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