Metabolic Engineering of Bacillus licheniformis for the Bioproduction of Nicotinamide Riboside from Nicotinamide and Glucose

Nicotinamide riboside (NR) is an important nucleotide, a precursor of nicotinamide adenine dinucleotide, that has attracted great attention as a promising nutraceutical to improve various symptoms of diabetes, vascular disease, and age-related physiological decline. Here, we systematically engineered Bacillus licheniformis to produce NR from glucose and nicotinamide (NAM) efficiently. Specifically, we selected the pathway using nicotinamide phosphoribosyltransferase (NAMPT) for converting NAM to nicotinamide mononucleotide (NMN) in the presence of 5-phosphoribosyl-1-pyrophosphate (PRPP) and increased the titer of NMN by engineering NAMPT and strengthening the PRPP supply. To prevent the degradation of NR, genes deoD and pupG coding purine nucleoside phosphorylase were deleted, resulting in 1.76 g/L NR accumulation in the culture medium. Then, we further deleted the nicotinamidase PncA encoding gene to prevent the synthesis of the byproduct nicotinate and overexpressed nucleotidase YfkN during the conversion of NMN to NR, where the titer of NR was increased by 226 and 41%, respectively. Notably, the efflux pump MdtL from Escherichia coli was first proven to be beneficial for NR production. Finally, combined with process optimization, the recombinant strain NR17 produced 11.33 g/L NR with a yield of 0.91 mol/mol NAM and a productivity of 0.44 g/(L·h) in a shake flask. An NR producer was constructed in this study and will be promising for the low-cost, high-quality industrial production of NR.