High-yield extraction and purification of glutathione reductase from baker's yeast.
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 40 degrees C. After removal of the toluene, the cells were then allowed to autolysis in buffer for 72 h at 4 degrees C. The cells were collected and resuspended in buffer. A second stage autolysis was carried out for another 96 h at 4 degrees 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.
Tsai
Y C
YC
Institute of Biochemistry, National Yang-Ming Medical College, Taiwan, R.O.C.
Yang
T Y
TY
Cheng
S W
SW
Li
S N
SN
Wang
Y J
YJ
D001329:Autolysis; D002846:Chromatography, Affinity; D004591:Electrophoresis, Polyacrylamide Gel; D005980:Glutathione Reductase / Q000302:isolation & purification*; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000201:enzymology*; D012685:Sepharose / Q000737:chemistry; D014050:Toluene / Q000494:pharmacology; D014227:Triazines / Q000737:chemistry
D014227:Triazines; C073381:Yellow H-E4G; D014050:Toluene; D012685:Sepharose; D005980:Glutathione Reductase