人CYP2D6基因体外表达体系和功能研究方法的建立  

Expressing in Vitro and Functional Study of Human CYP2D6 Gene

作　　者：吴振强[1,2] 秦胜营 

机构地区：[1]上海交通大学Bio-X研究院 [2]上海交通大学生命科学技术学院,中国上海200030

出　　处：《湖南师范大学自然科学学报》2013年第5期70-74,共5页Journal of Natural Science of Hunan Normal University

基　　金：国家863计划资助项目(2012AA02A515);国家973计划资助项目(2010CB529600);国家关键技术研究资助项目(2012BAI01B09)

摘　　要：根据GeneBank数据库中人CYP2D6基因构建表达载体pMA91-CYP2D6,转化酵母细胞AH22在体外进行表达,抽提所表达的CYP2D6微粒体蛋白进行Western Blot验证表明表达成功.该微粒体蛋白与探针药物异喹胍反应,利用HPLC检测生成产物4-羟基异喹胍,通过分析其得到酶动力学参数K m=10.14±0.94μmol/L,最终建立起完整的人CYP2D6表达体系和功能研究方法,为未来测定CYP2D6各等位基因对应酶的活性,实现临床个性化用药奠定基础.According to the GeneBank databases of human CYP2D6 gene, expression vectors pMA91- CYP2D6 were constructed by transforming the yeast cell AH22 to express CYP2D6 protein, and Western Blot resuits verify that the expression was successful in yeast cells. The microsomal protein was reacted with probe drugdebrisoquine to produce product 4-hydroxy debrisoquine, and its enzyme kinetic parameter Km, = 10.14 ± 0.94 μmol/L was obtained. Ultimately CYP2D6 expression system and functional study methods were established. This work would be a useful basis for evaluating the enzyme activity of CYP2D6 alleles and clinical personalized medicine.

关 键 词：CYP2D6 表达载体 微粒体蛋白 KM值 

分 类 号：Q786[生物学—分子生物学]