Abstract
Glutathione reductase was purified from chicken liver and some characteristics of the enzyme were investigated. The purification procedure was composed of four steps: preparation of homogenate, ammonium sulfate precipitation, 2′,5′-ADP Sepharose 4B affinity chromatography, and Sephadex G-200 gel filtration chromatography. Owing to the four consecutive procedures, the enzyme was purified 1714-fold, with a yield of 38%. Specific activity at the final step was 120 enzyme unit (EU)/mg of protein. The purified enzyme showed a single band on sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The molecular weight of the enzyme was found to be 100 kDa by Sephadex G-200 gel filtration chromatography, and the subunit molecular weight was found to be 43 kDa by SDS-PAGE. Optimum pH, stable pH, optimum ionic strength, and optimum temperature were 7.0, 7.4, 0.75 M Tris-HCl buffer including 1 mM EDTA, and 50°C, respectively. K M and V max values for NADPH and glutathione disulfide (GSSG) substrates were also determined for the enzyme.
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Erat, M., Demir, H. & Šakiroglu, H. Purification of glutathione reductase from chicken liver and investigation of kinetic properties. Appl Biochem Biotechnol 125, 127–138 (2005). https://doi.org/10.1385/ABAB:125:2:127
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DOI: https://doi.org/10.1385/ABAB:125:2:127