Feng Rongchen, Yan Ziting, Wei Guoguang, Wu Chaoqiang, Chen Feifei, Zhang Alei, Xu Sheng, Wang Xin, Chen Kequan
State Key Laboratory of Materials-Oriented Chemical Engineering, College of Biotechnology and Pharmaceutical Engineering, Nanjing Tech University, Nanjing, Jiangsu 211816, China.
College of Chemical Engineering, Inner Mongolia University of Technology, Hohhot, Inner Mongolia 010051, China.
Enzyme Microb Technol. 2025 Aug;188:110633. doi: 10.1016/j.enzmictec.2025.110633. Epub 2025 Mar 21.
Nicotinamide mononucleotide (NMN) is a pivotal molecule within the realm of metabolic health, serving as a precursor to nicotinamide adenine dinucleotide (NAD), a critical coenzyme in cellular energy metabolism. In recent years, the biological production of NMN has garnered significant interest. In this study, we developed the novel NRK-dependent synthesis routes for NMN production. Two strategies were designed to supply D-ribose-1-phosphate (R-1-P): (1) phosphorylation of exogenous D-ribose to ribose-5-phosphate (R-5-P) using engineered ribokinase (RK), followed by isomerization to R-1-P; (2) R-5-P synthesis from glucose through the pentose phosphate pathway. An optimized in vitro multi-enzyme cascade (XapA/PNP/NRK, PPM, NRK) identified NRK as the most efficient catalyst for NMN biosynthesis from D-ribose and niacinamide. In Escherichia coli, overexpression of this cascade, knockout of competing pathways, and secretion enhancement via a pelB signal peptide-fused PnuC transporter achieved an NMN titer of 62.0 mg L¹ .This work provides a viable alternative for the biosynthesis of NMN.
烟酰胺单核苷酸（NMN）是代谢健康领域中的关键分子，作为烟酰胺腺嘌呤二核苷酸（NAD）的前体，NAD是细胞能量代谢中的关键辅酶。近年来，NMN的生物生产引起了广泛关注。在本研究中，我们开发了用于NMN生产的新型NRK依赖性合成途径。设计了两种策略来供应D-核糖-1-磷酸（R-1-P）：（1）使用工程化核糖激酶（RK）将外源D-核糖磷酸化为核糖-5-磷酸（R-5-P），然后异构化为R-1-P；（2）通过戊糖磷酸途径从葡萄糖合成R-5-P。优化的体外多酶级联反应（XapA/PNP/NRK、PPM、NRK）确定NRK是从D-核糖和烟酰胺生物合成NMN的最有效催化剂。在大肠杆菌中，该级联反应的过表达、竞争途径的敲除以及通过pelB信号肽融合的PnuC转运蛋白增强分泌，实现了62.0 mg L⁻¹的NMN产量。这项工作为NMN的生物合成提供了一种可行的替代方法。
