title: Biochemical studies on glutathione S-transferase from the bovine filarial worm Setaria digitata.
abstract: Setaria digitata is a filarial worm of the cattle used as a model system for antifilarial drug screening, due to its similarity to the human filarial parasites Wuchereria bancrofti and Brugia malayi. Since filarial glutathione S-transferase (GST) is a good biochemical target for antifilarial drug development, a study has been undertaken for the biochemical characterization of GST from S. digitata. Cytosolic fraction was separated from the crude S.digitata worm homogenate by ultracentrifugation at 100,000 g and subjected to ammonium sulfate precipitation followed by affinity chromatography using GSH-agarose column. The kinetic parameters K (m) and V (max) values with respect to GSH were 0.45 mM and 0.105 μmol min(-1) mL(-1) respectively. With respect to 1-chloro-2,4-dinitrobenzene, the K (m) and V (max) values were 1.21 and 0.117 μmol min(-1) mL(-1) respectively. The effect of temperature and pH on GST enzyme activity was studied. The protein retained its enzyme activity between 0°C and 40°C, beyond which it showed a decreasing tendency, and at 80°C, the activity was lost completely. The enzyme activity was varying with change in pH, and the maximum GST activity was observed at pH 7.5. Gel filtration chromatographic studies indicated that the protein has a native molecular mass of about 54 kDa. The single band of GST subunit appeared in sodium dodecyl sulfate polyacrylamide gel electrophoresis was found to have molecular mass of ∼27 kDa. This shows that cytosolic S. digitata GST protein is homodimeric in nature.
authors.affiliation: Vector Control Research Centre (Indian Council of Medical Research), Indira Nagar, Pondicherry 605006, India.
authors.forename: Lakshmy
authors.initials: L
authors.lastname: Srinivasan
authors.forename: Nisha
authors.initials: N
authors.lastname: Mathew
authors.forename: Twinkle
authors.initials: T
authors.lastname: Karunan
authors.forename: Kalyanasundaram
authors.initials: K
authors.lastname: Muthuswamy
mesh_terms: D000818:Animals; D002417:Cattle; D005591:Chemical Fractionation; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D004137:Dinitrochlorobenzene / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D005260:Female; D005370:Filarioidea / Q000201:enzymology* / Q000302:isolation & purification; D005978:Glutathione / Q000378:metabolism*; D005982:Glutathione Transferase / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008970:Molecular Weight; D055503:Protein Multimerization; D013696:Temperature
chemical_list: D004137:Dinitrochlorobenzene; D005982:Glutathione Transferase; D005978:Glutathione