High-Yield Extraction and Purification of Glutathione Reductase from Baker's Yeast |
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Authors: | Ying-Chieh Tsai Tzu-Yu Yang Shiow-Wen Cheng Shui-Nin Li Yng-Jiin Wang |
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Institution: | Institute of Biochemistry, National Yang-Ming Medical College , Taipei, Taiwan, R. O. C. |
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Abstract: | Abstract Glutathione reductase was extracted from toluene-treated baker's yeast cells by a two-stage buffer autolysis method. The yeast cells were treated with toluene for 1 h at 40ddot]C. After removal of the toluene, the cells were then allowed to autolysis in buffer for 72 h at 4ddot]C. The cells were collected and resus-pended in buffer. A second stage autolysis was carried out for another 96 h at 4ddot]C. The enzyme was purified to 786-fold from the second stage cell autolysate by using two steps of affinity chromatography with triazine dyes (Yellow H-E4G and Yellow H-E6G) coupled to Sepharose CL-4B. By using this simplified method, 1.44 mg (165 units/mg) of glutathione reductase was obtained from 65 g (wet weight) of yeast cells, equivalent to 80% enzyme recovery. |
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