Enhancing nicotinamide mononucleotide production from glucose in Escherichia coli by genetic engineering

Abstract

Nicotinamide mononucleotide (NMN), as a healthcare product, plays positive effects on the human body. In this study, an engineered Escherichia coli (E. coli) strain was constructed to efficiently produce NMN from glucose and nicotinamide (NAM) through whole-cell biotransformation. First, a highly active nicotinamide phosphoribosyltransferase (NAMPT/NadV) from Vibrio phage KVP40 was screened, which showed the best productivity in catalyzing phosphoribosyl pyrophosphate (PRPP) and NAM to NMN in E. coli. Next, enzymatic activity of E. coli ribose-phosphate diphosphokinase (EcPRS) was improved and its expression level was optimized, which increased the supply of precursor PRPP. The results showed that mutant EcPRS^D115S had optimal activity and that the promoter and RBS combination (P_J23119 and RBS_III) optimized for EcPRS^D115S further accumulated NMN. With the systematic modification, such as the deletion of pfkA and pncC genes, and the expression of Cgzwf ^A243T and Cggnd ^S361F genes from Corynebacterium glutamicum, the biosynthesis of NMN was improved. Finally, NMN production was further increased after the introduction of NAM and NMN transporters. The strain ZY17 achieved NMN production of 1.61 ± 0.02 g/L in the shake flask level (OD600 = 10), and 13.3 ± 0.35 g/L in the 2-L bioreactor with a productivity of 1.11 g/(L.h) and an NAM conversion (α) of 85.3%. In conclusion, we propose a strategy that can effectively enhance the production of NMN in E. coli, which may bring new ideas for the biosynthesis of other nucleotide compounds.

Funding

the Green Bio-fabrication Program of China ,(No. 2021YFC2100900)