title: Production of phenylpyruvic acid from L-phenylalanine using an L-amino acid deaminase from Proteus mirabilis: comparison of enzymatic and whole-cell biotransformation approaches.
abstract: Phenylpyruvic acid (PPA) is an important organic acid that has a wide range of applications. In this study, the membrane-bound L-amino acid deaminase (L-AAD) gene from Proteus mirabilis KCTC 2566 was expressed in Escherichia coli BL21(DE3) and then the L-AAD was purified. After that, we used the purified enzyme and the recombinant E. coli whole-cell biocatalyst to produce PPA via a one-step biotransformation from L-phenylalanine. L-AAD was solubilized from the membrane and purified 52-fold with an overall yield of 13 %, which corresponded to a specific activity of 0.94 ± 0.01 μmol PPA min(-1)·mg(-1). Then, the biotransformation conditions for the pure enzyme and the whole-cell biocatalyst were optimized. The maximal production was 2.6 ± 0.1 g·L(-1) (specific activity of 1.02 ± 0.02 μmol PPA min(-1)·mg(-1) protein, 86.7 ± 5 % mass conversion rate, and 1.04 g·L(-1)·h(-1) productivity) and 3.3 ± 0.2 g L(-1) (specific activity of 0.013 ± 0.003 μmol PPA min(-1)·mg(-1) protein, 82.5 ± 4 % mass conversion rate, and 0.55 g·L(-1)·h(-1) productivity) for the pure enzyme and whole-cell biocatalyst, respectively. Comparative studies of the enzymatic and whole-cell biotransformation were performed in terms of specific activity, production, conversion, productivity, stability, need of external cofactors, and recycling. We have developed two eco-friendly and efficient approaches for PPA production. The strategy described herein may aid the biotransformational synthesis of other α-keto acids from their corresponding amino acids.
authors.affiliation: Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, Jiangnan University, Wuxi, 214122, China.|Synergetic Innovation of Center of Food Safety and Nutrition, Jiangnan University, Wuxi, 214122, China.
authors.forename: Ying
authors.initials: Y
authors.lastname: Hou
authors.affiliation: Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, Jiangnan University, Wuxi, 214122, China.|Synergetic Innovation of Center of Food Safety and Nutrition, Jiangnan University, Wuxi, 214122, China.
authors.forename: Gazi Sakir
authors.initials: GS
authors.lastname: Hossain
authors.affiliation: Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, Jiangnan University, Wuxi, 214122, China.|Synergetic Innovation of Center of Food Safety and Nutrition, Jiangnan University, Wuxi, 214122, China.
authors.forename: Jianghua
authors.initials: J
authors.lastname: Li
authors.affiliation: School of Chemical and Biomolecular Engineering, Georgia Institute of Technology, Atlanta, 30332, USA.
authors.forename: Hyun-Dong
authors.initials: HD
authors.lastname: Shin
authors.affiliation: Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, Jiangnan University, Wuxi, 214122, China. longliu@jiangnan.edu.cn.|Synergetic Innovation of Center of Food Safety and Nutrition, Jiangnan University, Wuxi, 214122, China. longliu@jiangnan.edu.cn.
authors.forename: Long
authors.initials: L
authors.lastname: Liu
authors.affiliation: Key Laboratory of Carbohydrate Chemistry and Biotechnology, Ministry of Education, Jiangnan University, Wuxi, 214122, China. gcdu@jiangnan.edu.cn.|Synergetic Innovation of Center of Food Safety and Nutrition, Jiangnan University, Wuxi, 214122, China. gcdu@jiangnan.edu.cn.
authors.forename: Guocheng
authors.initials: G
authors.lastname: Du
mesh_terms: D000596:Amino Acids / Q000378:metabolism*; D000642:Ammonia-Lyases / Q000235:genetics / Q000378:metabolism*; D001711:Biotransformation; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D015870:Gene Expression; D010649:Phenylalanine / Q000378:metabolism*; D010667:Phenylpyruvic Acids / Q000378:metabolism*; D011513:Proteus mirabilis / Q000201:enzymology* / Q000235:genetics; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism
chemical_list: D000596:Amino Acids; D010667:Phenylpyruvic Acids; D011994:Recombinant Proteins; D010649:Phenylalanine; D000642:Ammonia-Lyases; C031606:phenylpyruvic acid
keywords: L-Amino acid deaminase; Phenylpyruvic acid; Proteus mirabilis; Whole-cell biotransformation