Work

Systematic engineering for high-level production of β-nicotinamide mononucleotide from NAM and ribose

Year: 2024

Type: article

Abstract: β-nicotinamide mononucleotide (β-NMN) is a bioactive nucleotide that indirectly affects various cellular metabolic pathways, and has tremendous potential in the food, pharmaceutical, and cosmetics industries. In the present study, a multi-enzymatic cascade biocatalyst for the efficient synthesis of β-NMN was established. First, the optimal ribokinase gene was selected to construct a biocatalyst in a lower-endotoxin-level Escherichia coli strain to produce NMN from D-ribose. After fine-tuning the gene expression of the three cascade enzymes (RK/PRS/NAMPT), the production of NMN improved to 3.14 g/L. Next, the functions of membrane transporters that import NAM and export NMN were verified for NMN biosynthesis. Through the introduction of an ATP regeneration system, a drastic improvement in NMN biosynthesis at a yield of 8.74 g/L was achieved. The degradation of NMN and NAM in the bioconversion system was further inhibited after systematic modification of the related metabolic pathways. Finally, by incorporating two more copies of the key enzyme NAMPT and one copy of pyrophosphatase involved in inorganic pyrophosphate hydrolysis into the genome of the biocatalyst, the titer of NMN reached 12.2 g/L, with a molar conversion ratio of 98.7% from nicotinamide. This result provides an efficient and economical approach for producing NMN on an industrial scale. • A high-yield NMN biocatalyst produced 12.2 g/L NMN with a 1.02 g/L/h productivity. • Integration of RK and NAMPT genes into genome effectively improved NMN biosynthesis. • Introduction of an ATP regeneration system significantly increased NMN production.

Source: Food Bioscience

DOI: 10.1016/j.fbio.2024.105725