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.

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Acknowledgments

This work was financially supported by the Enterprise-university-research prospective program Jiangsu Province (BY2013015-37) and 863 Program (2014AA021200, 2014AA021201).

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There is no conflict of interest.

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Correspondence to Long Liu or Guocheng Du.

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Hou, Y., Hossain, G.S., Li, J. et al. Production of phenylpyruvic acid from l-phenylalanine using an l-amino acid deaminase from Proteus mirabilis: comparison of enzymatic and whole-cell biotransformation approaches. Appl Microbiol Biotechnol 99, 8391–8402 (2015). https://doi.org/10.1007/s00253-015-6757-0

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Keywords

  • l-Amino acid deaminase
  • Phenylpyruvic acid
  • Proteus mirabilis
  • Whole-cell biotransformation