Investigation of the effect of some antioxidant compounds on glutathione reductase (EC 1.8.1.7) enzyme

Glutathione reductase (GR, EC 1.8.1.7) is a specific antioxidant enzyme that catalyzes oxidized glutathione (GSSG) to reduced glutathione (GSH). GR enzyme maintains the cellular reduced GSH level and plays an central role in cell defense against reactive oxygen species. Glutathione reductase enzyme was purified from human erythrocyte by affinity chromatography in one step and the purification rate of GR was 6224 fold and specific activity was calculated as 9.586 EU/mg protein. The molecular weight of GR was determined to be 53 kDa by SDS-PAGE. The effect of thymoquinone and lycopene compounds on the GR activity purified from human erythrocytes was investigated. Both compounds showed inhibitory effect on GR activity. For thymoquinone and lycopene, % Activity vs. Inhibitor concentration was plotted. IC50 values for thymokinone and lycopene were calculated as 62.12 µM and 35.79 µM, respectively. Inhibition type and Ki values were determined from Lineveawer-Burk graph. The type of inhibition for thymoquinone and lycopene was found to be non-competitive inhibition. Ki value was calculated as 57.71 µM for thymokinone and 46.65 µM for lycopene. Keywords: Antioxidant, Glutathione reductase (GR), Inhibition, Lycopene, Purification, Thymoquinone.