Enhanced production of nicotinamide mononucleotide by high cell density culture of engineered Escherichia coli

Nicotinamide mononucleotide (NMN), an intermediate product of NAD+ biosynthesis, is commonly found intracellularly in living species. Recently, NMN has attracted great interest for the treatment of different diseases. Here, we developed a high cell density fed-batch culture system to produce NMN at higher concentrations using an engineered E. coil strain. Fed-batch culture using linear feeding strategy exhibited 9.38 ± 0.8, 14.8 ± 0.56, and 5.04 ± 0.21 g/L NMN concentrations, and 59.5 ± 0.44, 68.8 ± 0.49, and 36.2 ± 0.32 g/L dry cell weights, using different concentrations of nicotinamide (NAM) of 3, 5, and 10 g/L, respectively. A maximum NMN concentration of 17.2 ± 1.3 g/L with a productivity of 0.956 g/L/h was achieved using two intermittent additions of 5 g/L NAM after 6 h and 17 h. NMN obtained from the fermentation broth was purified using size exclusion chromatography, generating a yield of 53.9% NMN. Liquid chromatography-mass spectrometry and 1H nuclear magnetic resonance analysis was additionally used to validate the purity of NMN after purification. The fed-batch fermentation process applied in this study, using cheaper feed-stocks, demonstrated a remarkable enhancement in the production of therapeutically important NMN, making this a promising strategy for industrial-scale production of NMN.