Liu Yiming, Du Shanshan, Zhang Xiangxiang, Li Chao, Li Shubin, Xu Wenxia, Zhao Jingjing, Mu Wei, Han Xiaojun
State Key Laboratory of Urban Water Resource and Environment, MIIT Key Laboratory of Critical Materials Technology for New Energy Conversion and Storage, School of Chemistry and Chemical Engineering, Harbin Institute of Technology Harbin 150001 China
Chem Sci. 2025 Jun 6. doi: 10.1039/d5sc00852b.
Reduced nicotinamide adenine dinucleotide (NADH) salvage pathway reconstitution is a crucial step toward autonomous artificial cells. In living systems, d-ribose is a fundamental precursor intricately involved in the synthesis of nucleotides, and other critical metabolic pathways. An NADH synthesis pathway in artificial cells starting from d-ribose was constructed with a five-enzyme cascade containing ribokinase, ribose-phosphate pyrophosphokinase, nicotinamide phosphoribosyltransferase, nicotinamide mononucleotide adenylyltransferase, and formate dehydrogenase (RK, RPPK, NAMPT, NMNAT, and FDH), which efficiently converted 10 mM d-ribose into 415 μM NADH within 80 minutes under optimized conditions. The produced NADH was further used to drive the amino acid metabolism, , to convert NH and α-ketoglutarate to glutamate by introducing additional glutamate dehydrogenase (GDH) inside artificial cells. The successful reconstitution of the NADH synthesis pathway lays the foundation for fabricating artificial cells with complicated metabolic networks.
还原型烟酰胺腺嘌呤二核苷酸（NADH）补救途径的重建是迈向自主人工细胞的关键一步。在生命系统中，d-核糖是一种基本前体，复杂地参与核苷酸和其他关键代谢途径的合成。利用包含核糖激酶、磷酸核糖焦磷酸激酶、烟酰胺磷酸核糖转移酶、烟酰胺单核苷酸腺苷酸转移酶和甲酸脱氢酶（RK、RPPK、NAMPT、NMNAT和FDH）的五酶级联构建了一条从d-核糖开始的人工细胞NADH合成途径，在优化条件下，该途径能在80分钟内将10 mM d-核糖高效转化为415 μM NADH。所产生的NADH进一步用于驱动氨基酸代谢，即通过在人工细胞内引入额外的谷氨酸脱氢酶（GDH）将NH₃和α-酮戊二酸转化为谷氨酸。NADH合成途径的成功重建为制造具有复杂代谢网络的人工细胞奠定了基础。
