棘孢木霉几丁质酶基因的原核表达、复性、纯化以及酶学性质研究  

Expression in Escherichia coli,Purification,Renaturation and Characterization of Chitinase Gene from Trichoderma asperelluma

作　　者：丛大鹏[1] 李雅华[1] 咸洪泉[1] 

机构地区：[1]青岛农业大学生命科学学院,山东青岛266109

出　　处：《中国农学通报》2012年第11期34-38,共5页Chinese Agricultural Science Bulletin

基　　金：山东省自然科学基金"生防木霉菌的分子标记技术研究"(ZR2010CM040);青岛农业大学高层次人才启动基金"木霉菌几丁质酶新基因克隆表达及酶学性质分析"(631108)

摘　　要：为高效表达几丁质酶基因tachi1,研究Tachi1的酶学性质。采用PCR技术从棘孢木霉中克隆了几丁质酶基因tachi1并与pEHisTEV原核表达载体融合,转化大肠杆菌BL21。经异丙基-β-D-硫代半乳糖苷(IPTG)诱导,获得了以包涵体形式高效表达的Tachi1。包涵体经洗涤、溶解、复性和纯化后获得了高纯度的Tachi1。复性后的Tachi1具有较高的几丁质酶活性,该几丁质酶反应的最适温度为50℃,最适pH5.5。几丁质酶在30～35℃下稳定,在pH3～7之间几丁质酶有较高活性。The tachi1 gene of Trichoderma chitinase by Trichoderma asperelluma with pEHisTEV prokaryotic expression vector fusion was highly expressed in E.coli BL21 strains.After induced by isopropyl-β-D-thiogalactoside （IPTG）,the recombinant protein Tachi1 was highly expressed in the for mofinclusion bodies.The protein Tachi1 in the inclusion bodies could be solubilized,renatured and purified by a serial of treatments,including washing,denaturing and renaturing as well as purification.The renaturation protein possessed chitinase activity.

关 键 词：棘孢木霉 几丁质酶基因 表达 原核表达 蛋白质纯化 酶学性质 

分 类 号：Q936[生物学—微生物学]