乳酸杆菌中单宁酶基因的克隆、表达与纯化  

Cloning,expression and purification of tannase from Lactobacillus plantarum

作　　者：吴明波[1] 彭晓红[1] 温华[1] 任斌[1] 

机构地区：[1]四川大学纳米生物医学技术与膜生物学研究所,成都610041

出　　处：《四川大学学报（自然科学版）》2012年第2期479-483,共5页Journal of Sichuan University(Natural Science Edition)

基　　金：四川大学"985平台"建设资金

摘　　要：利用基因工程的方法从乳酸杆菌(Lactobacillus plantarum ATCC14917T)中克隆单宁酶基因,构建重组表达载体pNIC28-Bsa4-tanLpl,转化大肠杆菌(BL21-DE3)进行表达,并对单宁酶进行了纯化及简单的性质测定.亲和层析纯化后单宁酶产量大约为80 mg/L菌液,SDS-PAGE分析在50 kD处有单一的目的条带,用五倍子酸甲酯作为酶的底物对单宁酶性质的研究显示该酶在温度为30℃,pH为8的条件下活性最高(390 U/mg).A Tannase tanLpl （1410 bp） from Lactobacillus plantarum ATCC14917^T was cloned into the prokaryotic plasmid pNIC28-Bsa4. The recombinant protein was expressed in E. coli BL21-DE3 strain and purified through nickle affinity chromatography column with a production rate of 80mg/L. SDSPAGE analysis gave a molecular weight of approximately 50 kDa for a subunit of the protein. Subsequent enzymatic characterization with methyl gallate as a substrate revealed that tannase was most active at 30 ℃ at pH8 with a measured activity of 390 U/mg.

关 键 词：基因工程 单宁酶 LIC-PCR 五倍子酸 

分 类 号：Q81[生物学—生物工程]