Biosynthesis of a Therapeutically Important Nicotinamide Mononucleotide through a Phosphoribosyl Pyrophosphate Synthetase 1 and 2 Engineered Strain of Escherichia coli,ACS Synthetic Biology

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Biosynthesis of a Therapeutically Important Nicotinamide Mononucleotide through a Phosphoribosyl Pyrophosphate Synthetase 1 and 2 Engineered Strain of Escherichia coli
ACS Synthetic Biology ( IF 3.7 ) Pub Date : 2021-11-08 , DOI: 10.1021/acssynbio.1c00333
Anoth Maharjan ₁ , Mamata Singhvi ₁ , Beom Soo Kim ₁

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Nicotinamide mononucleotide (NMN), a precursor of NAD⁺, can be synthesized by the conversion of nicotinamide with the help of nicotinamide phosphoribosyl transferase (NAMPT) via the salvage pathway. NMN has recently gained great attention as an excellent therapeutic option due to its long-term effective pharmacological activities. In this study, we constructed a recombinant strain of Escherichia coli by inserting NAMPT and phosphoribosyl pyrophosphate synthetase 1 (PRPS1) and PRPS2 (from Homo sapiens) genes to investigate the effect of PRPS1 and PRPS2 on NMN synthesis. The metabolically engineered strain of E. coli BL21 (DE3) exhibited 1.57 mM NMN production in the presence of Mg²⁺ and phosphates in batch fermentation studies. For further improvement in NMN production levels, effects of different variables were studied using a response surface methodology approach. A significant increment was achieved with a maximum of 2.31 mM NMN production when supplemented with 1% ribose, 1 mM Mg²⁺ and phosphate, and 0.5% nicotinamide in the presence of a lactose (1%) inducer. Additionally, insertion of the PRPS1 and PRPS2 genes in the phosphoribosyl pyrophosphate synthesis pathway and individual gene expression studies facilitated a higher NMN production at the intracellular level than the reported studies. The strain exhibited intracellular production of NMN from cheap substrates such as glucose, lactose, and nicotinamide. Hence, the overall optimized process can be further scaled up for the economical production of NMN using a recombinant strain of E. coli BL21 (DE3), which is the future perspective of the current study.

中文翻译：

通过磷酸核糖焦磷酸合成酶 1 和 2 工程大肠杆菌菌株生物合成具有治疗意义的烟酰胺单核苷酸

烟酰胺单核苷酸 (NMN) 是 NAD ⁺的前体，可在烟酰胺磷酸核糖基转移酶 (NAMPT) 的帮助下通过补救途径转化烟酰胺来合成。由于其长期有效的药理活性，NMN 作为一种极好的治疗选择最近受到了极大的关注。在这项研究中，我们通过插入NAMPT和磷酸核糖焦磷酸合成酶 1 ( PRPS1 ) 和PRPS2 (来自智人) 基因构建了大肠杆菌的重组菌株，以研究PRPS1和PRPS2对 NMN 合成的影响。大肠杆菌的代谢工程菌株在分批发酵研究中，在存在 Mg ²⁺和磷酸盐的情况下，BL21 (DE3) 的 NMN 产量为 1.57 mM 。为了进一步提高 NMN 生产水平，使用响应面方法研究了不同变量的影响。^{在乳糖 (1%) 诱导剂存在的情况下，当添加 1% 核糖、1 mM Mg 2+}和磷酸盐以及 0.5% 烟酰胺时，NMN 产量会显着增加，最大产量为 2.31 mM 。此外，插入PRPS1和PRPS2与报道的研究相比，磷酸核糖焦磷酸合成途径中的基因和单个基因表达研究促进了细胞内水平的 NMN 产生。该菌株表现出从葡萄糖、乳糖和烟酰胺等廉价底物在细胞内产生 NMN。因此，可以进一步扩大整体优化过程，以使用重组大肠杆菌BL21 (DE3) 菌株经济地生产 NMN，这是当前研究的未来前景。

更新日期：2021-11-19

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