Oxidative Stress Response in Yeast: Purification and Characterization of Glutathione Reductase from Hansenula mrakii

• Published in: Bioscience, biotechnology, and biochemistry biotechnology, and biochemistry, 1996, Vol.60 (7), p.1207-1209
• Main Authors: Takeo, Miki, Yoshiyuki, Tsujimoto, Shinji, Miyabe, Kei-ichi, Sugiyama, Shingo, Izawa, Yoshiharu, Inoue, Akira, Kimura
• Format: Article
• Language: English
• Subjects: Biological and medical sciences; Biotechnology; Chloromercuribenzoates - pharmacology; Chromatography, DEAE-Cellulose; Fermentation; Fundamental and applied biological sciences. Psychology; Glutathione - metabolism; glutathione peroxidase; glutathione recycling; glutathione reductase; Glutathione Reductase - antagonists & inhibitors; Glutathione Reductase - isolation & purification; Glutathione Reductase - metabolism; Growth, nutrition, metabolism, transports, enzymes. Molecular biology; Hansenula mrakii; Hydrogen-Ion Concentration; Kinetics; Metals - pharmacology; Microbiology; Molecular Weight; Mycology; oxidative stress; Oxidative Stress - physiology; p-Chloromercuribenzoic Acid; Pichia - enzymology; Pichia - metabolism; Sulfhydryl Reagents - pharmacology
• ISSN: 0916-8451; 1347-6947
• DOI: 10.1271/bbb.60.1207

Glutathione reductase was purified from a yeast, Hansenula mrakii IFO 0895, to approximately 3500-fold with 59% activity yield. The enzyme was homogenous on polyacrylamide gel electrophoresis. The molecular weight of the enzyme was estimated to be 56 kDa by SDS-polyacrylamide gel electrophoresis, and 123 kDa by gel filtration using a calibrated Sephadex G-150 column. The K m values for glutathione disulfide and NADPH were 21.3μM and 14.3 μM, respectively. The enzyme was most active at pH 7.5, 55°C. The enzyme was stable up to 40°C, and between pHs 4 and 10. The enzyme was inhibited by p-chloromercuribenzoate and metal ions such as Fe 3+ , Cd 2+ , Cu 2+ and Zn 2+ .