Enhancement of biocatalytic efficiency by increasing substrate loading: enzymatic preparation of L-homophenylalanine.

Jielin Zhang, Tianyi Zhu, Xuri Wu, Yijun Chen

文献索引：Appl. Microbiol. Biotechnol. 97(19) , 8487-94, (2013)

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摘要

Enantiomerically pure L-homophenylalanine (L-HPA) is a key building block for the synthesis of angiotensin-converting enzyme inhibitors and other chiral pharmaceuticals. Among the processes developed for the L-HPA production, biocatalytic synthesis employing phenylalanine dehydrogenase has been proven as the most promising route. However, similar to other dehydrogenase-catalyzed reactions, the viability of this process is markedly affected by insufficient substrate loading and high costs of the indispensable cofactors. In the present work, a highly efficient and economic biocatalytic process for L-HPA was established by coupling genetically modified phenylalanine dehydrogenase and formate dehydrogenase. Combination of fed-batch substrate addition and a continuous product removal greatly increased substrate loading and cofactor utilization. After systemic optimization, 40 g (0.22 mol) of keto acid substrate was transformed to L-HPA within 24 h and a total of 0.2 mM NAD(+) was reused effectively in eight cycles of fed-batch operation, consequently giving an average substrate concentration of 510 mM and a productivity of 84.1 g l(-1) day(-1) for L-HPA. The present study provides an efficient and feasible enzymatic process for the production of L-HPA and a general solution for the increase of substrate loading.