State Key Laboratory of Food Science and Technology, Jiangnan University, Wuxi, 214122, China.
School of Life Sciences and Health Engineering, Jiangnan University, Wuxi, 214122, China.
Angew Chem Int Ed Engl. 2022 Sep 5;61(36):e202207077. doi: 10.1002/anie.202207077. Epub 2022 Jul 26.
L-Homophenylalanine (L-HPA) is a vital building block for the synthesis of numerous chiral drugs. However, the high cost of starting materials limits the industrial production of L-HPA. In this study, an enzymatic-spontaneous chemical cascade route for L-HPA production was designed based on retrosynthetic analysis. This route, using simple benzaldehyde and pyruvate as starting materials, is extremely cost-effective. The enzymes were screened and further assembled in E. coli, and TipheDH was identified as the rate-limiting enzyme. Therefore, TipheDH was engineered to improve its specific activity (by 82 %) and expression level (by 254 %), thus generating the best strain (W14). W14 exhibited the optimum enzyme activity ratio (1.7 : 1.1 : 1 : 1.8) and demonstrated production of 100.9 g L of L-HPA (with 94 % conversion, >99 % ee) in a 5-L reactor. This route effectively exploits the power of cascades and offers insight into avenues for synthesizing other valuable chemicals from inexpensive building blocks.
L-对羟基苯丙氨酸（L-HPA）是合成许多手性药物的重要基石。然而，起始原料的高成本限制了 L-HPA 的工业生产。在这项研究中，基于反合成分析，设计了一种酶促自发化学级联途径来生产 L-HPA。该途径使用简单的苯甲醛和丙酮酸作为起始原料，极具成本效益。筛选了酶并在大肠杆菌中进一步组装，鉴定出 TipheDH 是限速酶。因此，对 TipheDH 进行了工程改造以提高其比活性（提高 82%）和表达水平（提高 254%），从而生成最佳菌株（W14）。W14 表现出最佳的酶活比（1.7:1.1:1:1.8），并在 5-L 反应器中生产出 100.9 g/L 的 L-HPA（转化率>99%，ee 值>99%）。该途径有效地利用了级联的力量，并为从廉价原料合成其他有价值的化学品提供了思路。
