Characterization of a peroxidase in excretory-secretory product of adult Paragonimus westermani

Abstract

When activity of peroxidase in adult Paragonimus westermani was monitored using o-dianisidine and H2O2 as substrates, its specific activity was 1.5 times higher in excretory-secretory product (ESP) than in crude extract. The enzyme was purified by two purification steps of Sephacryl S-300 Superfine gel permeation and DEAE-Trisacryl M anion exchange chromatographies. Its activity increased 16.9 fold with 32.3% recovery. The enzyme was inhibited totally by 1 millimoles of dithiothreitol (DTT), 2-mercaptoethanol and azide. Molecular mass was 16 kDa in reducing SDS-polyacrylamide gel electrophoresis (PAGE) or 19 kDa in TSK-Blue gel filtration high performance liquid chromatography (HPLC), respectively. Special staining for peroxidase by diaminobenzidine on SDS-PAGE confirmed the activity. The peroxidase was less reactive to a paragonimiasis serum when observed by SDS-PAGE/immunoblot. In addition, specific activities of superoxide dismutase (SOD) and catalase were also identified in the ESP. High activities of these antioxidant enzymes in ESP indicate that they are parts of defense mechanisms against reactive oxygen intermediates from host.

Molecular cloning and characterization of a major egg antigen in Paragonimus westermani and its use in ELISA for the immunodiagnosis of paragonimiasis

“…A variety of diagnostic ELISA-based methods have been developed to improve test specificity. They include ELISAs based on species-specific monoclonal antibodies or the use of excretory/secretory antigens rather than a crude somatic extract (Chung et al 1993;Ichikawa and Ikeda 1995;Yong et al 1993). A cysteine proteinase antigen was reported to show higher specificity than crude adult worm extract or excretory/ secretory antigen (Ichikawa and Ikeda 1995;Ikeda 1998).…”