Purification and characterization of glutathione peroxidase from camel liver cytosol

ABSTRACT

Glutathione peroxidase [GSH H2O2 oxidoreductase, EC 1.11.1.9] was purified to homogeneity from camel liver cytosol by ammonium sulfate fractionation and chromatography on CM-Sephadex C-50, DEAE-Sephadex A-50, followed by Sephadex G-200 and Sephadex G-100. The purified enzyme was homogeneous as judged by SDS-PAGE. The molecular weight of the enzyme as determined by SDS-PAGE was 66 +/- 2 kDa and that by gel filtration was comparable, indicated that the enzyme protein was a single polypeptide. The enzyme selenium content was calculated to be 4.4 g atoms/mole enzyme protein. The basic and acidic amino acids represent 40% of the total protein content. Optimum pH was 8.4 using tris-HCl buffer at 37C. Optimum temperature was 55C and the Km values for both GSH and cumene-OOH were 0.8 and 1.6 mM, respectively. Cadmium ions [II] was found to be a potent inhibitor. The purified enzyme was stable for more than 2 months under frozen conditions in the presence of dithiothreitol [DTT]