A recombinant thioredoxin-glutathione reductase from Fasciola hepatica induces a protective response in rabbits |
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Authors: | Maggioli Gabriela Silveira Fernando Martín-Alonso José M Salinas Gustavo Carmona Carlos Parra Francisco |
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Institution: | aDepartamento de Bioquímica y Biología Molecular, Instituto Universitario de Biotecnología de Asturias, Universidad de Oviedo, 33006 Oviedo, Spain;bUnidad de Biología Parasitaria, Facultad de Ciencias, Instituto de Higiene, Av. A. Navarro 3051, CP 11600 Montevideo, Uruguay;cCátedra de Inmunología, Facultad de Química, Instituto de Higiene, Av. A. Navarro 3051, CP 11600 Montevideo, Uruguay |
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Abstract: | Antioxidant systems are fundamental components of host–parasite interactions, and often play a key role in parasite survival. Here, we report the cloning, heterologous expression, and characterization of a thioredoxin glutathione reductase (TGR) from Fasciola hepatica. The deduced polypeptide sequence of the cloned open reading frame (ORF) confirmed the experimental N-terminus previously determined for a native F. hepatica TGR showing thioredoxin reductase (TR) activity. The sequence revealed the presence of a fusion between a glutaredoxin (Grx) and a TR domain, similar to that previously reported in Schistosoma mansoni and Echinococcus granulosus. The F. hepatica TGR sequence included an additional redox active center (ACUG; U being selenocysteine) located at the C-terminus. The addition of a recombinant selenocysteine insertion sequence (SECIS) element in the Escherichia coli expression vector, or the substitution of the native selenocysteine by a cysteine, indicated the relevance of this unusual amino acid residue for the activity of F. hepatica TGR. Rabbit vaccination with recombinant F. hepatica TGR reduced the worm burden by 96.7% following experimental infection, further supporting the relevance of TGR as a promising target for anti Fasciola treatments. |
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Keywords: | Abbreviations: DTNB 5 5&prime -dithiobis(2-nitrobenzoic acid) EST expressed sequence tag FAD flavin adenine dinucleotide GR glutathione reductase Grx glutaredoxin GSH reduced glutathione GSSG oxidized glutathione HED β-hydroxyethyl disulfide IPTG d-thiogalactopyranoside" target="_blank">isopropyl β-d-thiogalactopyranoside ORF open reading frame PBS phosphate-buffered saline SECIS selenocysteine insertion sequence TGR thioredoxin glutathione reductase TR thioredoxin reductase Trx thioredoxin |
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