高山被孢霉中异柠檬酸脱氢酶的异源表达及酶学特性的研究
Heterologous Expression and Characterization Isocitrate Dehydrogenase from the Oleaginous Fungus Mortierella alpina
异柠檬酸脱氢酶（Isocitrate dehydrogenase，IDH）是一类在能量代谢、氨基酸合成和维生素合成中起重要作用的酶。为探究高山被孢霉（Mortierella alpina）中NADP+依赖型异柠檬酸脱氢酶（NADP+-MaIDH）的酶学性质，通过构建重组质粒pET28a-MaIDH1转入大肠杆菌（Escherichia coli）BL21（DE3）中异源表达NADP+-MaIDH1，以获取目的蛋白。研究结果表明，NADP+-MaIDH1在大肠杆菌BL21（DE3）中得到成功诱导表达，其比酶活为571.7nmol min-1 （mg 蛋白）-1，为阴性对照菌株的8.2倍。NADP+-MaIDH1的最适温度和最适pH分别为35℃和pH 8.0，该酶在温度低于40℃时具有较好的热稳定性，高于40℃时酶的活性迅速降低；在pH 6.5-8.0的范围内具有较好的pH稳定性，pH值高于8.0时酶的活性迅速降低。这为深入研究NADP+-MaIDH1的功能提供了一定的指导意义。
Isocitrate dehydrogenase (IDH) plays an important role in energy metabolism, amino acid synthesis and vitamin synthesis. In order to investigate the enzymatic properties of NADP+-dependent isocitrate dehydrogenase in Mortierella alpina (NADP+-MaIDH), the recombinant plasmid pET28a-MaIDH was transformed into Escherichia coli BL21 (DE3). The results showed that NADP+-MaIDH was successfully expressed in E. coli BL21 (DE3). The activity of NADP+-MaIDH was up to 571.7 nmol min-1 (mg protein)-1, which was about 8.2 times of the control strain. The optimum temperature and pH of NADP+-MaIDH was 35 C and 8.0, respectively. The recombinant enzyme had good thermal stability when the temperature was lower than 40 C, and the activity of the enzyme decreased rapidly when it was higher than 40 C. The NADP+-MaIDH can maintain high activity at the pH value raging from 6.5 to 8.0 and the activity of the enzyme sharply decreasesd when the pH was higher than 8.0. This provided theoretical guidance for further study of the function of the isocitrate dehydrogenase.
张灏、唐鑫、孙晓琪、陈卫、陈海琴
生物化学生物工程学
生物工程异柠檬酸脱氢酶高山被孢霉大肠杆菌酶学性质
BioengineeringIsocitrate dehydrogenaseMortierella alpinaEscherichia coliCharacterization
张灏,唐鑫,孙晓琪,陈卫,陈海琴.高山被孢霉中异柠檬酸脱氢酶的异源表达及酶学特性的研究[EB/OL].(2019-02-03)[2025-08-23].http://www.paper.edu.cn/releasepaper/content/201902-11.点此复制
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