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.