Identification and localization of glutathione S-transferase as a potential target enzyme in Brugia species

Abstract

Brugia filarial nematodes are pathogenic lymphatic-dwelling parasites that, like other helminths, may modify the host's defense mechanisms by a major detoxification process involving glutathione-binding proteins such as glutathione S-transferases (GSTs). In the present study, soluble extracts of third-stage larvae, adult male and female worms, microfilariae of either B. pahangi or B. malayi or the adult worm excretory-secretory products of B. malayi were used to determine GST activity. These extracts and affinity-purified fractions of B. pahangi adult worms had a specific enzymatic activity when 1-chloro-2,4-dinitrobenzene was used as a substrate. The observance of this enzyme in all life cycle stages of Brugia spp. demonstrates its ubiquitous nature. Lavage of intraperitoneally infected jirds, but not that of uninfected jirds, also showed increased enzymatic activity, suggesting that GST is secreted in vivo. Soluble proteins of both Brugia spp. were strongly recognized by antibodies in sera from rabbits immunized with affinity-purified native GST of Onchocerca volvulus. Immunohistochemical studies localized these proteins in adult worms, demonstrating cross-reactivity between the GST of these two filarial nematodes. The effect of this enzyme on the motility and viability of adult worms, microfilariae, and larvae was tested in vitro using a battery of known GST inhibitors. Of all those tested, ethacrynic acid, N-ethylmalemide, 4-nitropyridine-oxide, or 1-chloro-2,4-dinitrobenzene at micromolar concentrations reduced the viability and motility of microfilariae, third-stage larvae, and adult worms. These results suggest that Brugia GSTs are major metabolic enzymes and may play an important role in the parasite's survival.

Structural Analysis and Antibody Response to the Extracellular GlutathioneS-Transferases fromOnchocerca volvulus

“…4 for GP1 [44][45][46][47][48][49][50][51][52][53][54][55][56][57] and GP4 [144][145][146][147][148][149][150][151][152][153][154][155][156] . The assignment and the ratios of the glycan structures found at individual glycosylation sites of OvGST1a are as follows: for GP1 [44][45][46][47][48][49][50][51][52][53][54][55][56][57] and the relative ratios of oligosaccharides linked to glycosylation sites 1 (N50), 2 (N79), and 3 (N134) appear to be rather similar, at glycosylation site 4 (N144) clearly larger structures, bearing Man 5 GlcNAc 2 to Man 9 GlcNAc 2 , were detected. For further characterization of the (glyco)peptides, individual fractions from another HPLC run were collected and the amino acid sequences of all glycopeptides were confirmed by Edman sequencing.…”

Biochemical Characterization and Vaccine Potential of a Heme-Binding Glutathione Transferase from the Adult Hookworm Ancylostoma caninum

“…This suggests that Ac-gst-1 mRNA might be transcribed but not translated in the non-blood-feeding stages or expressed at such a low level that it cannot be detected with antibody. Helminth GSTs have been reported from every developmental stage (31,39,40); however, one helminth GST from the lung fluke, Paragonimus westermani, was expressed only by adult flukes (19). The adult-specific expression of Ac-GST-1 protein suggests that the protein may play an important role in the survival of the adult worm in the host during blood feeding.…”

Novel Phage Display-Based Subtractive Screening To Identify Vaccine Candidates ofBrugia malayi

“…Using these approaches several potential vaccine candidates have been identified and reported to have varying degrees of protection in animal models (1,20,24,28,29,33,37,50). Although there are effective drugs available for the control of filariasis, developing a vaccine remains a promising strategy for mass control of this mosquito-borne infection in areas of endemicity (24,29,28,50).…”