Abstract:In order to reduce the cost and simplify the production process of β-nicotinamide mononucleotide (β-NMN) synthesis by enzyme catalytic method, on the basis of the previously established system of β-NMN synthesis by six enzymes, the six enzymes were combined in pairs to construct a double-enzyme co-expression recombinant strain, and the crude enzyme solution obtained by co-expression was used for multi-enzyme catalytic cascade reaction to synthesize β-NMN. Finally, the yield of β-NMN was as high as 72.90% when the substrate D-ribose concentration was 10 mmol/L. In addition, we optimized the induction temperature and IPTG induction concentration of the recombinant strains to further improve the expression of the recombinant enzymes. Finally, we obtained salt ions and polyhydroxy compounds that significantly improved the stability of 5-phosphate ribose-1-pyrophosphate synthase (EcPrs). The effect of stabilizers on the enzymatic preparation of β-NMN was further investigated. Among them, Mn2+, lactose and sorbitol had no effect on the yield of β-NMN. In this study, the establishment of cascade catalytic reaction for the preparation of β-NMN under dual-enzyme co-expression system and the optimization of EcPrs stability provide a new idea for the enzymatic synthesis of β-NMN.
