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食品与发酵工业  2021, Vol. 47 Issue (12): 1-9    DOI: 10.13995/j.cnki.11-1802/ts.026161
  研究报告 本期目录 | 过刊浏览 | 高级检索 |
代谢工程改造大肠杆菌合成L-组氨酸
李梦莹1,2, 吕雪芹1,2, 刘延峰1,2, 李江华1,2, 堵国成1,2, 吴剑荣2*, 刘龙1,2*
1(江南大学 未来食品科学中心,江苏 无锡,214122)
2(糖化学与生物技术教育部重点实验室(江南大学),江苏 无锡,214122)
Metabolic engineering of Escherichia coli for increased synthesis of L-histidine
LI Mengying1,2, LYU Xueqin1,2, LIU Yanfeng1,2, LI Jianghua1,2, DU Guocheng1,2, WU Jianrong2*, LIU Long1,2*
1(Science Center for Future Foods, Jiangnan University, Wuxi 214122, China)
2(Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, Jiangnan University, Wuxi 214122, China)
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摘要 该研究主要通过多种代谢工程策略对大肠杆菌进行改造从而增强菌株L-组氨酸的合成。首先,用人工的开放阅读框(hisL'-λattB'-trpE)替换大肠杆菌(Escherichia coli,E.coli) BL21(DE3)L-组氨酸操纵子的前导区,同时表达来源于E.coli的ATP转磷酸核糖基酶突变体基因hisGE271K后,菌株L-组氨酸产量达到872.50 mg/L;其次,通过比较来自谷氨酸棒状杆菌(Corynebacterium glutamicum,C.glutamicum)ATCC130132、粘质沙雷氏菌(Serratia marcescens,S.marcescens)ZJZ626和E.coli BL21(DE3)的11种hisG在不同抑制物浓度下的酶活力以及过表达11种hisG基因时L-组氨酸的产量,筛选出最优hisG基因,L-组氨酸产量提高至1 480.42 mg/L;然后,通过CRISPR/Cas9技术将最优hisG整合至基因组;进一步比较不同来源的Prs对菌株L-组氨酸产量的影响,从中筛选出较优Prs进行基因组整合,结果显示,摇瓶发酵72 h后,菌株L-组氨酸产量提高至3 898.06 mg/L,96 h时产量提高至5 574.63 mg/L。该研究为将来L-组氨酸的工业化生产奠定了良好的基础。
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李梦莹
吕雪芹
刘延峰
李江华
堵国成
吴剑荣
刘龙
关键词:  L-组氨酸  大肠杆菌  ATP转磷酸核糖基酶  代谢工程  反馈抑制    
Abstract: In this study, several metabolic engineering strategies were used to improve the synthesis of L-histidine in Escherichia coli. Firstly, the precursor region of L-histidine operon in E. coli was replaced by an artificial open reading frame (hisL'-λattB'-trpE), and the titer of L-histidine was increased to 872.50 mg/L after the expression of hisGE271K. Secondly, the enzyme activities was compared under different concentrations of inhibitors, and L-histidine titer was analyzed when 11 kinds of hisG from Corynebacterium glutamicum ATCC 130132, Serratia marcescens ZJZ626 and E. coli BL21 (DE3) were expressed. The optimal hisG was selected and L-histidine titer was increased to 1 480.42 mg/L. Next, CRISPR/Cas9 was used to integrate the optimal hisG in the genome. Finally, after comparing the effects of two different Prs on L-histidine production, the better one was selected and integrated in the genome, resulting in the L-histidine titer of 3 898.06 mg/L in 72 h and 5 574.63 mg/L in 96 h. This work laid a good foundation for the industrial production of L-histidine in the future.
Key words:  L-histidine    Escherichia coli    ATP transphosphatase    metabolic engineering    feedback inhibition
收稿日期:  2020-11-12      修回日期:  2020-12-16           出版日期:  2021-06-25      发布日期:  2021-07-22      期的出版日期:  2021-06-25
基金资助: 国家重点研发计划项目(2020YFA0908300);国家自然基金面上项目(21676119)
作者简介:  硕士研究生(吴剑荣副教授和刘龙教授为共同通讯作者,E-mail:kinowu@jiangnan.edu.cn;longliu@jiangnan.edu.cn)
引用本文:    
李梦莹,吕雪芹,刘延峰,等. 代谢工程改造大肠杆菌合成L-组氨酸[J]. 食品与发酵工业, 2021, 47(12): 1-9.
LI Mengying,LYU Xueqin,LIU Yanfeng,et al. Metabolic engineering of Escherichia coli for increased synthesis of L-histidine[J]. Food and Fermentation Industries, 2021, 47(12): 1-9.
链接本文:  
http://sf1970.cnif.cn/CN/10.13995/j.cnki.11-1802/ts.026161  或          http://sf1970.cnif.cn/CN/Y2021/V47/I12/1
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