Identification of GR and TrxR systems in Setaria cervi: Purification and characterization of glutathione reductase.
Parasitol Int
; 62(2): 193-8, 2013 Apr.
Article
en En
| MEDLINE
| ID: mdl-23305756
ABSTRACT
The glutathione reductase (GR) and thioredoxin reductase (TrxR) are important enzymes of the redox system that aid parasites to maintain an adequate intracellular redox environment. In the present study, the enzyme activity of GR and TrxR was investigated in Setaria cervi (S. cervi). Significant activity of both enzymes was detected in the somatic extract of adult and microfilariae stages of S. cervi. Both GR and TrxR were separated by partial purification using ammonium sulfate fractionation and DEAE ion exchange chromatography suggesting the presence of both glutathione and thioredoxin systems in S. cervi. The enzyme glutathione reductase (ScGR) was purified to homogeneity using affinity and ion exchange chromatography that resulted in 90 fold purification with a yield of 11.54%. The specific activity of the ScGR was 643U/mg that migrated as a single band on SDS-PAGE. The subunit molecular mass was determined to be ~50kDa while the optimum pH and temperature were found to be 7.0 and 35°C respectively. The activation energy (Ea) was calculated from the slope of Arrhenius plot as 16.29±1.40kcal/mol. The Km and Vmax were determined to be 0.27±0.045mM; 30.30±1.30U/ml with NADPH and 0.59±0.060mM; 4.16±0.095U/ml with GSSG respectively. DHBA, a specific inhibitor for GR has completely inhibited the enzyme activity at 1µM concentration. The inhibition of ScGR activity with NAI (IC50 0.71mM), NEM (IC50 0.50mM) and DEPC (IC50 0.27mM) suggested the presence of tyrosine, cysteine and histidine residues at its active site. Further studies on characterization and understanding of these antioxidant enzymes may lead to designing of an effective drug against lymphatic filariasis.
Texto completo:
1
Colección:
01-internacional
Banco de datos:
MEDLINE
Asunto principal:
Reductasa de Tiorredoxina-Disulfuro
/
Setaria (Nematodo)
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Filariasis
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Glutatión Reductasa
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Estadios del Ciclo de Vida
Tipo de estudio:
Diagnostic_studies
/
Prognostic_studies
Límite:
Animals
Idioma:
En
Revista:
Parasitol Int
Asunto de la revista:
PARASITOLOGIA
Año:
2013
Tipo del documento:
Article
País de afiliación:
India