Key Laboratory of Industrial Biotechnology Ministry of Education and School of Biotechnology, Jiangnan University, 1800 Lihu Avenue, Wuxi 214122, China; State Key Laboratory of Food Science and Resources, Jiangnan University, 1800 Lihu Avenue, Wuxi 214122, China.
Zhengzhou Tobacco Research Institute of CNTC, Zhengzhou 450001, China.
Enzyme Microb Technol. 2024 Oct;180:110482. doi: 10.1016/j.enzmictec.2024.110482. Epub 2024 Jul 20.
β-nicotinamide mononucleotide (β-NMN) is a key precursor of nicotinamide adenine dinucleotide, and becomes attractive in the nutrition and health care fields, but its enzymatic synthesis is expensive. In this study, a six-enzyme cascade catalytic system was constructed to produce β-NMN. Using D-ribose and nicotinamide as substrates, the β-NMN yield reached 97.5 % catalyzed by purified enzymes. Then, after knocking out the genes encoding proteins that consume β-NMN in E. coli BL21(DE3), the similar β-NMN yield, 97.2 %, using the crude enzymes could be also obtained. After that, β-NMN synthesis was performed under increased substrate concentration, and 'modular' crude enzymes cascade catalytic reaction system was proposed to reduce the inhibition of polyphosphate on ribose-phosphate diphosphokinase activity, and the β-NMN yield reached 78.4 % at 10 mM D-ribose, which is 1.82 times of that in 'one-pot' reaction and represents the highest β-NMN preparation level with phosphoribosylpyrophosphate as the core reported till now.
β-烟酰胺单核苷酸（β-NMN）是烟酰胺腺嘌呤二核苷酸的关键前体，在营养和保健领域具有吸引力，但酶法合成成本高昂。本研究构建了一个六酶级联催化体系来生产β-NMN。使用 D-核糖和烟酰胺作为底物，经纯化酶催化，β-NMN 的产率达到 97.5%。然后，在大肠杆菌 BL21(DE3)中敲除消耗 β-NMN 的蛋白编码基因后，使用粗酶也可以得到类似的β-NMN 产率，为 97.2%。之后，在提高底物浓度下进行 β-NMN 合成，并提出“模块”粗酶级联催化反应体系来降低多磷酸盐对核糖-磷酸二磷酸激酶活性的抑制作用，β-NMN 的产率在 10mM D-核糖时达到 78.4%，是“一锅法”反应的 1.82 倍，是目前为止以磷酸核糖焦磷酸为核心报道的最高β-NMN 制备水平。
