利用大肠杆菌tyrB-aspA基因串联表达制备L-苯丙氨酸  

L-phenylanine Production by the tyrB and aspA Co-expression in Escherichia coli

作　　者：范长胜[1] 李欣[1] 高山峨[1] 王海蛟[1] 焦晔[1] 王建刚[1] 梁国新[1] 陶菊红[1] 

机构地区：[1]复旦大学生命科学学院微生物系

出　　处：《食品与发酵工业》2005年第7期1-4,共4页Food and Fermentation Industries

基　　金：国家自然科学基金资助(No.30470045)项目

摘　　要：从大肠杆菌K12菌株中获得tyrB和aspA基因,将2个基因串联并克隆到pλPR质粒上,然后转化到E·coliP2392菌株进行表达。相应酶活力和氨基酸产量检测结果表明,单基因表达和双串联基因串联表达菌株的酶活性成倍提高,与原始菌株相比,AspA活力分别提高243%与239%,TyrB活力分别提高到247%和236%。以基因双串联表达菌株E·coliPBA的菌体悬浮液为酶原,以铵盐、反丁烯二酸和苯丙酮酸为酶转化底物,反应3h后,苯丙酮酸转化苯丙氨酸的效率为93%,苯丙氨酸产率达到0·6g/(g·h)(细胞),是对照菌株的4·7倍。TyrB encoding aromatic-amino-acid transaminase and aspA encoding aspartase were obtaineo from Escherichia coli K12. They were constructed into a vector pλPR in tandem. Then the recombinant vector was introduced into E. coli P2392 for the gene expression and enzymatic activity determination. AspA activity of the strain harboring pλPR-aspA and pλPR-tyrB-aspA has an increase of 243% and 239%. TyrB activity of the strain harboring pλPR-tyrB and pλPR-tyrB-aspA has an increase of 247% and 236% relative to that of the host strain respectively. When fumaric acid （PA）, NH4Cl and phenylpyruvic acid （PPA） were used as synthetic substrates in a reaction system for 3 h, E. coli PBA/pλPR-tyrB-aspA achieved a PPA conversion rate of 93% with a phenylalanine yield of 0.6g/g of cells·h^-1, which is 4.7-fold compare with the control strain E. coli P2392.

关 键 词：aspA tyrB 天冬氨酸酶 芳香氨基酸转氨酶 苯丙氨酸 L-苯丙氨酸 串联基因 串联表达 大肠杆菌 E.COLI 

分 类 号：TQ920[轻工技术与工程—发酵工程]