D-glucaric acid (GA) has been identified as among promising biotechnological alternatives to oil-based chemicals. GA and its derivatives are widely used in food additives, dietary supplements, drugs, detergents, corrosion inhibitors and biodegradable materials. The increasing availability of a GA market is improving the cost-effectiveness and efficiency of various biosynthetic pathways. In this study, an engineered Escherichia coli strain GA10 was constructed by systematic metabolic engineering. This involved redirecting metabolic flux into the GA biosynthetic pathways, blocking the conversion pathways of D-glucuronic acid (GlcA) and GA into by-products, introducing an in situ NAD+ regeneration system and fine-tuning the activity of the key enzyme, myo-inositol oxygenase (Miox). Subsequently, the culture medium was optimized to achieve the best performance of the GA10 strain. GA was produced at 5.35 g/L (extracellular and intracellular), with a maximized yield of ∼0.46 mol/mol on D-glucose and glycerol, by batch fermentation. This work demonstrates efficient biosynthetic pathways of GA in E. coli by metabolic engineering and should accelerate the application of GA biosynthetic pathways in industrial processes.

中文翻译：

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用于大肠杆菌代谢工程的组合合成途径微调和辅因子再生显着提高 D-葡糖二酸的产量

D-葡糖二酸 (GA) 已被确定为油基化学品的有前途的生物技术替代品之一。GA及其衍生物广泛用于食品添加剂、膳食补充剂、药物、洗涤剂、缓蚀剂和生物降解材料。GA 市场的日益普及正在提高各种生物合成途径的成本效益和效率。在这项研究中，通过系统代谢工程构建了工程大肠杆菌菌株 GA10。这包括将代谢通量重新定向到 GA 生物合成途径、阻断 D-葡萄糖醛酸 (GlcA) 和 GA 转化为副产物的途径、引入原位 NAD+ 再生系统并微调关键酶 myo-肌醇加氧酶（Miox）。随后，优化培养基以实现GA10菌株的最佳性能。通过分批发酵，GA 的产量为 5.35 g/L（细胞外和细胞内），在 D-葡萄糖和甘油上的最大产量约为 0.46 mol/mol。这项工作通过代谢工程证明了大肠杆菌中 GA 的有效生物合成途径，并应加速 GA 生物合成途径在工业过程中的应用。