# Advances in the Synthesis and Physiological Metabolic Regulation of Nicotinamide Mononucleotide
## 3 Biosynthesis of NMN
### Selection of NMN Synthesis Hosts  
Currently, _E. coli_ serves as the principal engineering strain for NMN biosynthesis research. As a prototypical model organism, _E. coli_ possesses a well-defined genetic background and a plethora of gene editing tools, facilitating metabolic engineering modifications. Additionally, there are reports of utilizing alternative host strains for NMN synthesis. The food-grade expression system utilizing lactic acid bacteria, known for its efficiency and safety, has gained extensive attention in the food fermentation industry. Leveraging lactic acid bacteria as the host cells for NMN synthesis has emerged as a promising strategy. [PERSON] et al. [70] screened three candidates from a library of 174 strains of lactic acid bacteria, among which _Lactobacillus_ RD011727 produced the highest NMN titer of 2 mg/L. [PERSON] et al. [71] used CRISPR/nCas9 gene editing technology to knock out the _nadR_ gene in _Lactococcus lactis_ NZ9000 and heterologously expressed the _nadE\({}^{*}\)_ gene from _Francisella tulareis_, resulting in a final intracellular NMN production of 2289 \(\upmu\)mol/L/mg from 10 g/L of externally added NAM in 12 h.  
Previously, NMN synthesis was conducted in microorganisms using glucose as the carbon source. As a non-food feedstock, methanol holds great promise for bio-manufacturing from non-sugar carbon sources. The conversion of captured carbon dioxide into methanol through hydrogenation catalysis, followed by microbial conversion, presents an attractive avenue for low-carbon biosynthesis [72]. In recent years, some progress has been made in the research on NMN synthesis using _P. pastoris_, which expanded the substrate spectrum of NMN biosynthesis. [PERSON] et al. [73] achieved secretory expression of the NRK enzyme in _P. pastoris_ cells using the pPIC9K plasmid vector, with a crude extracellular enzyme preparation achieving a molar conversion rate of 92.6%. [PERSON] et al. [74] heterologously expressed the NAMPT enzyme and introduced NiaP and PnuC transport proteins to enhance substrate uptake and product excretion in _P. pastoris_ cells, enabling whole-cell NMN synthesis and offering a new strategy for industrial production. [PERSON] et al. [75] constructed the _Pichia pastoris_ strain PC110-A0305 using the CRISPR-Cas9 gene editing system and heterologously expressed NAMPT in this strain for whole-cell biocatalytic synthesis of NMN. Methanol was added at 24 h intervals during the shake-flask fermentation stage to induce NMN synthesis. The intracellular NMN content reached a peak of 1.46 mg/g DCW after 24 h. Furthermore, the study established a high-fat diet (HFD) obesity model to investigate the anti-obesity effects of NMN. The results demonstrated that NMN supplementation modulated lipid metabolism, ameliorated obesity as well as lipid and glucose metabolism dysregulation, reduced hepatic fat accumulation, and mitigated liver injury in HFD mice. The study provides a theoretical basis for the production of NMN from methanol in _Pichia pastoris_, as well as its nutritional applications. Leveraging methanol as an abundant and cost-effective substrate, and _Pichia pastoris_ can be engineered as a cellular factory for producing pure NMN or diversified products rich in NMN, with significant economic and environmental implications.