碱性果胶酯裂解酶工程菌的构建  

Construction of Engineering Strain Producing Pectate Lyase

作　　者：诸葛斌[1] 堵国成[1] 沈微[1] 陈坚[1] 

机构地区：[1]江南大学生物工程学院环境生物技术研究室/工业生物技术教育部重点实验室,江苏无锡214036

出　　处：《应用与环境生物学报》2006年第4期547-549,共3页Chinese Journal of Applied and Environmental Biology

基　　金：国家"863"资助项目(2003AA322050)~~

摘　　要：从本研究室筛选的BacillussubtilisWSHB04-02菌株中扩增出编码碱性果胶酯裂解酶的结构基因PL,将其插入大肠杆菌分泌表达载体pET22b(+)多克隆位点,得到重组载体pET22b(+)PL.序列分析表明,所获PL基因与已报道的B.subtilisSO113的PL基因的同源性为98%.重组载体在大肠杆菌BL21(DE3)中得到表达.SDS-PAGE分析显示,表达产物的分子量(Mr)均为43×103,同核酸序列测定所推导的值相符.该工程菌经IPTG诱导后,胞内酶活为8U/mL.研究发现,碱性果胶酯裂解酶不仅在胞内有酶活,胞外也有酶活,说明所构建的表达体系可使该酶向胞外分泌.The structure gene PL from Bacillus subtilis WSHB04-02 encoding pectate lyase was amplified by PCR method. Comparison with the structure gene PL from B. subtilis SO113 revealed 98% identity. The pET22b （ ＋ ） vector harboring PL gene was transformed into E. coil BL21DE3. The PL was expressed in E. coil BL21 （DE3）. SDS - PAGE analysis showed that the molecular weight of expressed recombinant PL was about 43×10^3, which was the same as the calculated molecular weight. The intracellular pectate lyase activity of recombinant E. coil reached up to 8 U/mL under the induction of IPTG.

关 键 词：碱性果胶酯裂解酶 基因 表达 

分 类 号：TQ920[轻工技术与工程—发酵工程] Q789[生物学—分子生物学]