O-Succinyl-L-homoserine (OSH) is an important platform chemical in production of C₄ chemicals such as succinic acid, homoserine lactone, γ‑butyrolactone, and 1,4‑butanediol. The production of OSH through chemical method or the current engineering strain is difficult and not optimal, and thereby there remains a need to develop new engineering strategy. Here, we engineered an OSH overproducing Escherichia coli strain through deleting the degradation and competitive pathways, overexpressing thrA and metL to enhance the metabolic flux from asparate to homoserine. Additionally, increasing the precursor succinyl-CoA supply through simultaneously knocking out sucD and overexpressing sucA further increased the yield of OSH. The engineered strain OSH9/pTrc-metA11-yjeH with above strategies produced OSH at the concentration of 24.1 g/L (0.609 g/g glucose) in batch fermentation. To gain detailed insight into metabolism of the engineered strain, comparative metabolic profiling was performed between the engineered and wide-type strain. The metabolomics data deciphered that the carbon was directed toward the OSH biosynthesis resulting in less flexibility of the genetically modified strain than the wide-type strain.

O-琥珀酰-L-高丝氨酸（OSH）是生产琥珀酸、高丝氨酸内酯、γ-丁内酯和1,4-丁二醇等C ₄化学品的重要平台化学品。通过化学方法或目前的工程菌株生产 OSH 困难且不是最优的，因此仍然需要开发新的工程策略。在这里，我们通过删除降解和竞争途径、过表达thrA和metL来增强从天冬氨酸到高丝氨酸的代谢通量，设计了一种 OSH 过度生产的大肠杆菌菌株。此外，通过同时敲除sucD和过表达来增加前体琥珀酰辅酶 A 的供应sucA进一步提高了 OSH 的产量。具有上述策略的工程菌株 OSH9/pTrc- metA 11- yjeH在分批发酵中产生浓度为 24.1 g/L（0.609 g/g 葡萄糖）的 OSH。为了详细了解工程菌株的代谢，在工程菌株和宽型菌株之间进行了比较代谢分析。代谢组学数据破译了碳直接用于 OSH 生物合成，导致转基因菌株的灵活性低于宽型菌株。