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<p>DAAO is applied as a potential catalyst in the biosynthesis of L-PPT. However, its low solubility expression constrains its broader industrial application. Herein, a novel DAAO derived from <i>Cladophialophora</i> <i>carrionii</i> (<i>Cc</i>DAAO) was identified, which demonstrated superior catalytic performance toward D-Ala (specific activity: 106.38 ± 1.21&#xa0;U/mg, <i>K</i><sub>m</sub>: 1.56 ± 0.06&#xa0;mM), along with remarkable thermostability and broad substrate spectrum. Under optimal culture conditions, the soluble expression level of <i>Cc</i>DAAO was enhanced through a co-expression strategy with molecular chaperones, and the enzyme activity increased by 36.3% compared with the initial level. Subsequently, <i>Cc</i>DAAO was constructed as a fusion protein (CGD) with catalase from <i>Geobacillus</i> sp. CHB1 (<i>Gb</i>CAT) and applied in a D-amino acid aminotransferase (DAAT)-mediated cascade system. In a 2&#xa0;L reaction system, this cascade system achieved complete conversion (&gt; 99%) of 1&#xa0;M D,L-PPT within 8&#xa0;h, exhibiting a yield of 11.26&#xa0;g/L/h for PPO, which represents a significant improvement over existing reports. This study presents a promising practical approach for the industrial production of optically pure L-PPT.</p>

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Identification of a novel D-amino acid oxidase and its application in deracemization of D, L-phosphinothricin

  • 期刊论文

  • 发表日期：

    2025年8月9日

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  • Li-Qun Jin,
  • Meng-Dan Liu,
  • Zi-Yu Guan,
  • Yi-Xin Li,
  • Ya-Ping Xue,
  • Zhi-Qiang Liu,
  • Yu-Guo Zheng

摘要

DAAO is applied as a potential catalyst in the biosynthesis of L-PPT. However, its low solubility expression constrains its broader industrial application. Herein, a novel DAAO derived from Cladophialophora carrionii (CcDAAO) was identified, which demonstrated superior catalytic performance toward D-Ala (specific activity: 106.38 ± 1.21 U/mg, Km: 1.56 ± 0.06 mM), along with remarkable thermostability and broad substrate spectrum. Under optimal culture conditions, the soluble expression level of CcDAAO was enhanced through a co-expression strategy with molecular chaperones, and the enzyme activity increased by 36.3% compared with the initial level. Subsequently, CcDAAO was constructed as a fusion protein (CGD) with catalase from Geobacillus sp. CHB1 (GbCAT) and applied in a D-amino acid aminotransferase (DAAT)-mediated cascade system. In a 2 L reaction system, this cascade system achieved complete conversion (> 99%) of 1 M D,L-PPT within 8 h, exhibiting a yield of 11.26 g/L/h for PPO, which represents a significant improvement over existing reports. This study presents a promising practical approach for the industrial production of optically pure L-PPT.