Enhancing the biosynthesis of nicotinamide mononucleotide in Lactococcus lactis by heterologous expression of FtnadE.

BACKGROUND: Nicotinamide mononucleotide (NMN), a key intermediate of nicotinamide adenine dinucleotide, plays an important in anti-aging and disease. Lactococcus lactis, an important probiotic lactic acid bacteria (LAB), has shown great potential for the biosynthesis of NMN, which will significantly affect the probiotic effects of the dairy products. RESULTS: We used the CRISPR/nCas9 technique to knockout nadR gene of L. lactis NZ9000 to enhance the accumulation of NMN by 61%. The nadE* gene from Francisella tularensis with codon optimization was heterologous in L. lactis NZ9000ΔnadR and has a positive effect on NMN production. Combined with optimization of the concentration of substrate nicotinamide, a final intracellular NMN titer was 2289 µmol L-1  mg-1 with 10 g L-1 nicotinamide supplement, which was 5.7-fold higher than that of the control. The transcription levels of key genes (pncA, nadD and prs1) involved in NMN biosynthesis were up-regulated by more than two-fold, indicating that the increase of NMN titer was attributed to FtnadE* heterologous expression. CONCLUSION: Our study provides a better understanding of the NMN biosynthesis pathway in L. lactis, and can facilitate NMN production in LAB via synthetic biology approaches. © 2022 Society of Chemical Industry.

[Optimization of fermentation of recombinant human Endostatin (rh-Endostatin) expression in Escherichia coli].

The fermentation process of recombinant human Endostatin expression in Escherichia coli BL21 (DE3) was studied. The effects of factors such as concentration of IPTG, induction time, cultivation temperature and feeding strategies were investigated. Beside that, by changing the temperature to 40 degrees C after induction, the high-density cultivation finished in a much shorter period. After 9 hours cultivation, the optical density (OD) at 600 nm reached 140 and the yield of inclusion body was 3 g/L. While E. coli system was used, protein with better activity and stability was obtained. The cost was much lower and the producing process was much steadier. It will meet the demands of the industrial production.