产L-阿拉伯糖异构酶基因工程菌的构建及优化  

Construction and optimization of genetically engineered bacteria producing L-arabinose isomerase

作　　者：张艳芳 王晓茹 张一帆 张春枝[1] ZHANG Yanfang;WANG Xiaoru;ZHANG Yifan;ZHANG Chunzhi(School of Biological Engineering,Dalian Polytechnic University,Dalian 116034,China)

机构地区：[1]大连工业大学生物工程学院,辽宁大连116034

出　　处：《大连工业大学学报》2024年第4期257-260,共4页Journal of Dalian Polytechnic University

基　　金：辽宁省教育厅科学研究经费项目(面上项目)(LJKZ0514)。

摘　　要：L-阿拉伯糖异构酶(L-AI)可专一性的作用于底物D-半乳糖,生成D-塔格糖。为获得高产L-AI的菌株,将来源于短乳杆菌L.brevis sp. D-tag 1的L-AI编码基因araA基因连接到pET-30a(+)表达质粒中构建重组质粒,并将重组质粒通过热激法转入E.coli BL21(DE3),构建得到基因工程菌E.coli BL21(DE3)/pET-30a(+)-araA。根据宿主大肠杆菌的密码子偏好性,对araA基因进行密码子优化,并将优化的araA_o基因连接到pET-30a(+)表达质粒中构建重组质粒,采用相同的方法构建得到基因工程菌E.coli BL21(DE3)/pET-30a(+)-araA_(o)。结果表明,在相同培养条件下,优化后的菌株较未优化的表达量有明显提高,酶活提高了79.2%。L-arabinose isomerase(L-AI)can specifically act on the substrate D-galactose to produce D-tagatose.To obtain a high-yield L-AI strain,the L-AI coding gene araA gene derived from L.brevis sp.D-tag1 was connected to pET-30a(+)expression plasmid to construct a recombinant plasmid.Then the recombinant plasmid was transferred into E.coli BL21(DE3)by heat shock method,and E.coli BL21(DE3)/pET-30a(+)-araA was constructed.The codon optimization of araA gene was carried out according to the codon preference of host E.coil.The optimized araA o gene was connected to pET-30a(+),then transferred into E.coli BL21(DE3)to construct E.coli BL21(DE3)/pET-30a(+)-araA_(o).The results showed the optimized strain showed a significant increase in expression levels compared to the unoptimized strain,with an increase in enzyme activity of 79.2%.

关 键 词：L-阿拉伯糖异构酶 D-塔格糖 密码子优化 

分 类 号：Q939.97[生物学—微生物学]