Nicotinamide mononucleotide (NMN) is a derivative of the vitamin B group, which naturally exists in fruits, vegetables, and meat.It has been widely used in the field of food, health care products, and degenerative diseases treatment.Nicotinamide phosphoribosyltransferase (NAMPT) is a rate-limiting enzyme in the NMN salvage synthesis pathway.The best NAMPT in current research is VpNadV derived from Vibrio bacteriophage, which has several defects, such as poor activity and low substrate utilization.In addition, its crystal structure has not been fully resolved.Based on the NMN fluorescence reaction system, this study aimed to construct a high-throughput screening system, and then to screen highly active VpNadV mutants combined with enzyme engineering, finally to apply for efficient synthesis of NMN in Escherichia coli.Through the high-throughput screening and enzyme engineering, a mutant with high enzymatic activity, VpNadVL139V+R382G, was selected, which enzyme activity was 1.46-fold than that of the wild type.Based on the mutant, 3.68 g/L NMN was achieved in shake flasks, and 24.4 g/L NMN was obtained after scaling up in a 5 L fermenter.In this study, a high-throughput screening system for Nicotinamide phosphoribosyltransferase (NAMPT) mutants was constructed, and then the catalytic mechanism of VpNadV was briefly analyzed, and finally the positive effect of NMN synthesis by mutants was further verified by fermentation, providing ideas for the efficient production of NMN and nicotinamide derivatives.
LI Zhanyan
,
WANG Ke
,
ZENG Weizhu
,
ZHOU Jingwen
. Screening of highly active nicotinamide phosphoribosyltransferase to enhance nicotinamide mononucleotide synthesis[J]. Food and Fermentation Industries, 2025
, 51(22)
: 51
-59
.
DOI: 10.13995/j.cnki.11-1802/ts.041876
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