代谢工程改造大肠杆菌发酵生产β-烟酰胺单核苷酸

摘要/Abstract

摘要： 为了实现大肠杆菌发酵高效生产β-烟酰胺单核苷酸（β-NMN），设计模块化代谢改造策略。首先，对烟酰胺（NAM）和β-NMN支路代谢涉及的8个酶进行失活，减少底盘细胞对前体和产物的额外消耗。其次，通过引入NAM输入蛋白（BcNiaP）、β-NMN输出蛋白（BmPnuC）、PRPP合成酶（Prs）和烟酰胺磷酸核糖转移酶（Nampt），敲除调节蛋白PurR，工程菌N12’摇瓶发酵可积累0.34 g/L的β-NMN；此后，比对筛选发现Comamonadaceae bacterium来源的Nampt酶活性较高且对底盘细胞负担较小；通过进一步强化BmPnuC和Prs的表达水平，工程菌N18摇瓶发酵β-NMN产量提高至1.36 g/L。最后，利用发酵罐分批补料发酵38 h，β-NMN产量达到10.2 g/L，NAM到β-NMN的摩尔转化率为74.5%。研究构建的β-NMN发酵菌株具有遗传背景清晰、无营养缺陷、无需诱导等优势，工业前景良好。

关键词: β-烟酰胺单核苷酸, 大肠杆菌, 烟酰胺磷酸核糖转移酶, 代谢改造, 发酵

Abstract: In order to realize efficient the fermentative production of β-Nicotinamide mononucleotide (β-NMN) by Escherichia coli, the modular metabolic engineering strategies were designed. Firstly, eight enzymes involved in nicotinamide (NAM) and β-NMN pathway metabolism were inactivated to reduce the additional consumption of precursors and products by chassis cells. Secondly, by introducing BcNaiP, BmPnuC, Prs and Nampt, deleting regulatory protein PurR, the engineered strain N12' can accumulate 0.34 g/L β-NMN in shake-flask fermentation. Then, by comparison and screening, we found that the Nampt enzyme from Comamonadaceae bacteriumChitinphaga Pinensis had higher activity and less burden on the chassis cells. By further strengthening the expression level of BmPnuC and Prs, the β-NMN titer of the engineered strain N18 increased to 1.36 g/L in shake-flask fermentation. Finally, after fed-batch fermentation for 38 h, β-NMN titer reached 10.2 g/L, and the molar conversion rate of NAM to β-NMN was 74.5%. The β-NMN fermentation strain constructed in this study has the advantages of clear genetic background, no nutritional defects, no induction, and has a good industrial prospect.

Key words: β-Nicotinamide mononucleotide, Escherichia coli, Nicotinamide phosphoribosyl transferase, Metabolic modification, Fermentation

中图分类号:

Q939.97

安俊侠王倩倩王昭颖刘欢徐庆阳范晓光. 代谢工程改造大肠杆菌发酵生产β-烟酰胺单核苷酸[J]. 食品科学, 0, (): 0-0.

XU Qingyang. Metabolic Engineering of Escherichia coli for Fermentative Production of β-nicotinamide Mononucleotide[J]. FOOD SCIENCE, 0, (): 0-0.

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