菠萝泛菌木聚糖酶xynB的克隆表达与活性研究  

Cloning,Expression and Activity Analysis of xynB from Pantoea ananatis in Escherichia coli

作　　者：侯进慧[1] 王宾[1] 王富威[1] 

机构地区：[1]徐州工程学院食品(生物)工程学院,江苏徐州221008

出　　处：《生物技术》2013年第5期28-31,共4页Biotechnology

基　　金：江苏省高校自然科学研究项目(12KJD550003);徐州工程学院校级培育科研项目(XKY2011110);江苏省大学生实践创新项目资助

摘　　要：目的:克隆、表达菠萝泛菌(Pantoea ananatis)木聚糖酶基因。方法:以菠萝泛菌Y1菌株基因组DNA为模板,通过PCR方法获得该菌木聚糖酶基因XynB,将该基因与pET表达载体连接,转化E.coli BL21(DE3)菌株,构建表达重组木聚糖酶的菌株pETxynB/BL21(DE3)。结果:采用0.1mmol/L的IPTG诱导重组菌株,3h后表达出大量的重组木聚糖酶。采用保持蛋白活性的方法回收获得纯度大于95%的重组木聚糖酶。等电聚焦实验分析纯化后的重组木聚糖酶XynB等电点为7.3。该重组酶在60℃、pH 6时相对酶活性最高。结论:该文为菠萝泛菌木聚糖酶的进一步开发提供了研究基础。Objective：Cloning and expressing Pantoea ananatis xylanase gene. Method：With Pantoea ananatis Y1 genomic DNA as tem-plate, a xylanase gene, xynB, was cloned by the method of PCR. The gene was ligated with pET expression vector and transformed into E-coli BI21 （ DE3 ） to construct a recombinant xylanase strain pETxynB/BL21 （ DE3 ）. Result： After induced with 0. lmmol/L IPTG for 3hr, recombinant protein of XynB was successfully expressed by the recombinant strain. The recombinant protein was recovered by a retai-ning activity method, and purity of the recombinant protein is more than 95%. The result of isoelectric focusing shows, pI of the purified recombinant XynB is 7. 3. Relative activity of the recombinant enzyme reaches its peak at 60℃ and pH 6. Conclusion：The paper provided research basis for further study of Pantoea ananatis xylanase.

关 键 词：PANTOEA ananatis 木聚糖酶 ESCHERICHIA COLI 重组蛋白 

分 类 号：Q814[生物学—生物工程]