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: A modular metabolic engineering for Escherichia coli was designed for efficient production of β-nicotinamide mononucleotide (β-NMN). First, eight enzymes involved in the shunt metabolic pathways of nicotinamide (NAM) and β-NMN were inactivated to reduce the additional consumption of precursors and products by chassis cells. Second, by introducing BcNaiP, BmPnuC, 5-phosphoribosyl-1-pyrophosphate synthetase (Prs) and nicotinamide phosphoribosyl transferase (Nampt) and deleting the regulatory protein PurR, an engineered strain N12’ was obtained, which could accumulate 0.34 g/L β-NMN in shake-flask fermentation. Then, we found that the Nampt enzyme from Comamonadaceae bacterium had higher activity and caused less burden on the chassis cells. By further strengthening the expression levels of BmPnuC and Prs, the β-NMN titer of the engineered strain N18 was increased to 1.36 g/L in shake-flask fermentation. Finally, after fed-batch fermentation for 38 h, the β-NMN titer reached 10.2 g/L, and the molar conversion rate of NAM to β-NMN was 74.5%. This engineered strain has the advantages of clear genetic background, no nutritional defects, and no need for induction and therefore, has a good prospect for industrial application.

Key words: β-nicotinamide mononucleotide; Escherichia coli; nicotinamide phosphoribosyl transferase; metabolic modification; fermentation