南京师大学报（自然科学版）Issue(3)：95-99,105,6.
基于基因组的TEV蛋白酶的高表达
Chromosome-based TEV Protease Overexpression
摘要
Abstract
Due to its high proteinase cleavage activity,specificity and effective in a wide range of conditions,Tobacco etch virus protease ( TEV ) has many applications, ranging from protein isolation to proteomics study. Currently, TEV is produced by plasmid-based overexpression in Escherichia coli BL21(DE3). Yet,the method has inherent disadvantages, for example,it needs antibiotic to maintain the plasmid which may bring impurities during the protein purification;and non-homogeneity of the strain population which may reduce the yield. Herein,we report the recombineering mediated in-tegration of MBP fused TEV gene under the strong T7 promoter into E. coli BL21 ( DE3 ) chromosome. Intracellular digestion of chromosomal based overexpression of MBP-TEV released TEV which was subsequently isolated though Ni-NTA affinity purification, the yield of TEV was up to 4. 2 mg/L. Purified TEV shows fine protease activity. The engineered strain has the potential to be used for large scale TEV purification,the established recombineering method can be a platform for the genome engineering and heterologus gene expression in E. coli BL21(DE3).关键词
基于基因组/重组工程/TEV/蛋白表达Key words
chromosome-based/recombineering/TEV/protein expression分类
生物科学引用本文复制引用
李玲,凌文,石牡丹,尚广东..基于基因组的TEV蛋白酶的高表达[J].南京师大学报（自然科学版）,2014,(3):95-99,105,6.基金项目
国家自然科学基金(NSFC81273412) （NSFC81273412）
