Fukuda H, Moriguchi M, Tochikura T
J Biochem. 1980 Mar;87(3):841-6. doi: 10.1093/oxfordjournals.jbchem.a132814.
Glyoxylate reductase II was purified about 2,400-fold from a cell extract of baker's yeast by protamine sulfate treatment, and column chromatographies on DEAE-cellulose, hydroxylapatite, Sephadex G-150, and phosphocellulose. The purified enzyme was electrophoretically homogeneous. The molecular weight was determined to be approximately 65,000 by gel filtration. The enzyme was greatly stabilized by the addition of 20% (v/v) glycerol. It catalyzed the reduction of glyoxylate and hydroxypyruvate and was specific for NADPH as an electron donor, but showed slight affinity towards NADH. The Michaelis constants for glyoxylate, hydroxypyruvate, NADPH, and NADH were found to be 16mM, 1.4 mM, 5.7 microM, and 0.43 mM, respectively. The enzyme was inhibited by p-chloromercuribenzoate (PCMB) and iodoacetate, but inhibition was prevented by dithiothreitol (DTT) or L-cysteine. The reduction of glyoxylate and hydroxypyruvate was not stimulated by anions.
通过硫酸鱼精蛋白处理以及在DEAE - 纤维素、羟基磷灰石、葡聚糖G - 150和磷酸纤维素上的柱色谱法，从面包酵母的细胞提取物中纯化出乙醛酸还原酶II，纯化倍数约为2400倍。纯化后的酶在电泳上是均一的。通过凝胶过滤测定其分子量约为65000。加入20%（v/v）甘油可使该酶得到极大的稳定。它催化乙醛酸和羟基丙酮酸的还原反应，对作为电子供体的NADPH具有特异性，但对NADH有轻微亲和力。发现乙醛酸、羟基丙酮酸、NADPH和NADH的米氏常数分别为16mM、1.4 mM、5.7 microM和0.43 mM。该酶受到对氯汞苯甲酸（PCMB）和碘乙酸的抑制，但二硫苏糖醇（DTT）或L - 半胱氨酸可防止这种抑制。阴离子不会刺激乙醛酸和羟基丙酮酸的还原反应。
