植物乙醇酸氧化酶分离纯化方法的改进.pdfVIP

植物学通报 2006, 23 (6): 703~707 Chinese Bulletin of Botany . . 技术与方法 植物乙醇酸氧化酶分离纯化方法的改进 * 郭晋雅,董宇亮,杨净,徐杰 ,尹汗萍,曾秋莲 华南师范大学生命科学学院, 广东省植物发育生物工程重点实验室, 广州 510631 -1 . 摘要 通过缩短DEAE-Cellulose柱长度, 加快流速并采用pH8.8 的80 mmol L Tris-HCl为洗脱液, 可在9 -1 . 小时内快速地从菠菜、菜心和豆角绿叶中纯化得到乙醇酸氧化酶。该酶具高活性(54.6～197.0 U mg ) 及高等电点(pI 10.0) 。产率为4.1%～71.5%, 纯化倍数为21.6～122.68。经SDS检测均有40 kD带, 表明3种植物乙醇酸氧化酶的亚基大小无区别。 关键词 乙醇酸氧化酶, 纯化, 菠菜, 菜心, 豆角 Improvement of the Purification Method of Plant Glycolate Oxidase * Jinya Guo, Yuliang Dong, Jing Yang, Jie Xu , Hanping Yin, Qiulian Zeng College of Life Sciences, South China Normal University, Guangdong Key Lab of Biotechnology for Plant Development, Guangzhou 510631, China Abstract By reducing the length of diethylaminoethanol-cellulose and increasing the pH in elution solution, we developed an improved method for purifying glycolate oxidase with high isoelectric value (pI 10.0) from green leaves of Spinacia oleracea, Brassica parachinensis, and Phaseolus vulgaris . Purified glycolate oxidase -1 . showed high specific activity (54.6-197.0 U mg ) and high pI (10.0). We achieved 21.6-122.68-fold purifi- cation and 4.1%-71.5% yield from crude solution during this purification procedure. SDSof the