产长叶烯大肠杆菌工程菌的构建及发酵优化  

Construction of engineered Escherichia coli to produce longlifolene and optimization of the fermentation process

作　　者：夏芙蓉 魏丽娟 刘欢 巴利民 刘艳辉[1] XIA Furong;WEI Lijuan;LIU Huan;BA Limin;LIU Yanhui(College of Life Science and Technology,Beijing University of Chemical Technology,Beijing 100029,China;China Animal Husbandry Industry Co.,Ltd.,Beijing 100091,China)

机构地区：[1]北京化工大学生命科学与技术学院,北京100029 [2]中牧实业股份有限公司研究院,北京100091

出　　处：《生物加工过程》2025年第2期190-198,共9页Chinese Journal of Bioprocess Engineering

基　　金：国家自然科学基金青年项目(21606014)。

摘　　要：以大肠杆菌BL21(DE3)为底盘细胞,通过筛选不同来源的法尼基焦磷酸(FPP)合酶以及对其内源2甲基赤藓醇磷酸(MEP)途径中的关键酶基因1脱氧D木酮糖5磷酸合酶(dxs)、1脱氧D木酮糖5磷酸还原异构化酶(dxr)和异戊烯基焦磷酸异构酶(idi)进行过表达,构建出中心代谢途径优化的且具有合成长叶烯能力的大肠杆菌细胞工厂。为增强细胞工厂合成长叶烯的前体物供给,对发酵条件进行优化,筛选出最佳诱导剂异丙基βD硫代半乳糖苷(IPTG)浓度为0.1 mmol/L,诱导剂添加时间为0 h(即种子液接至发酵液时),诱导温度为30℃,最适培养基为TB培养基,最终摇瓶中长叶烯产量达到3.85 mg/L,较之前所报道的产量提高了45.8%,为进一步实现长叶烯的高产奠定了基础。In this study,Escherichia coli(E.coli)BL21(DE3)was used as the chassis cell.Farnesyl pyrophosphate(FPP)synthase from different sources were screened and key enzyme genes in the endogenous MEP pathway,including 1-deoxy-D-xylulose-5-phosphate synthase(dxs),1-deoxy-D-xylulose-5-phosphate reductoisomerase(dxr),isopentenyl pyrophosphate isomerase(idi)were overexpressed.An engineered E.coli cell factory with an optimized central carbon metabolic pathway capable of synthesizing longifolene efficiently was constructed.To further enhance the precursor supply for the synthesis of longleafene in the cell factory,fermentation conditions were optimized in this study.It was indicated that the optimal concentration of the inducer isopropylβ-D-thiogalactoside(IPTG)was 0.1 mmol/L,with the preferable addition time being 0 h(when the seed culture was transferred to the fermentation broth)and the optimal induction temperature at 30℃.The most suitable medium was TB medium.Finally,the yield of longifolene in the shake flask reached 3.85 mg/L,a 45.8%increase compared to the value reported in the literature.This study laid a good foundation for further research to realize the high yield of longifolene using E.coli.

关 键 词：MEP途径 长叶烯 大肠杆菌 发酵优化 

分 类 号：Q789[生物学—分子生物学]