title: Enhancement of biocatalytic efficiency by increasing substrate loading: enzymatic preparation of L-homophenylalanine.
abstract: 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.
authors.affiliation: Laboratory of Chemical Biology and State Key Laboratory of Natural Medicines, China Pharmaceutical University, 24 Tongjia St., Nanjing, Jiangsu Province, 210009, China.
authors.forename: Jielin
authors.initials: J
authors.lastname: Zhang
authors.forename: Tianyi
authors.initials: T
authors.lastname: Zhu
authors.forename: Xuri
authors.initials: X
authors.lastname: Wu
authors.forename: Yijun
authors.initials: Y
authors.lastname: Chen
mesh_terms: D000594:Amino Acid Oxidoreductases / Q000378:metabolism*; D000613:Aminobutyrates / Q000378:metabolism*; D001709:Biotechnology / Q000379:methods*; D005560:Formate Dehydrogenases / Q000378:metabolism*; D007651:Keto Acids / Q000378:metabolism; D009243:NAD / Q000378:metabolism
chemical_list: D000613:Aminobutyrates; D007651:Keto Acids; D009243:NAD; C014328:2-amino-4-phenylbutyric acid; D005560:Formate Dehydrogenases; D000594:Amino Acid Oxidoreductases; C022342:phenylalanine oxidase