Scholarlink

<p>Chiral amines, as essential chiral building blocks in drug synthesis, present a considerable challenge in biomanufacturing due to the requirement for highly efficient stereoselective synthesis. In this study, we successfully cloned and heterologously expressed a novel (<i>R</i>)-amine transaminase, MagAT, from <i>Mycolicibacterium agri</i>. Systematic analysis showed optimal activity at pH 7.0, with the highest reaction rate occurring within 30&#xa0;min at 50 ℃. However, considering overall thermal stability, 40℃ was selected as the operating temperature for subsequent experiments. Furthermore, the enzyme retained nearly 100% catalytic activity in the presence of 10% methanol, DMSO, and chloroform. Kinetic analysis demonstrated that MagAT possessed high substrate affinity, with Michaelis constants (K<sub>m</sub>) of 5.13 ± 0.45&#xa0;mM for (<i>R</i>)-1-phenylethan-1-amine and 3.73 ± 0.34&#xa0;mM for pyruvate. Substrate specificity studies indicated that MagAT efficiently catalyzed the conversion of C3-C8 aliphatic amines (e.g., (<i>R</i>)-2-aminooctane) and monocyclic aromatic amines (e.g., (<i>R</i>)-1-phenylethan-1-amine), whereas its catalytic activity significantly declined for sterically hindered substrates such as naphthylamine. 3D structural modeling with AlphaFold3 and molecular docking showed that steric hindrance from key residues in the O-pocket critically influences substrate specificity. This study not only provides an efficient biocatalyst for the biosynthesis of chiral amines but also establishes a theoretical basis for the rational design of transaminases with broader substrate adaptability.</p>

错误：搜索内容不能为空，请输入英文关键词
错误：关键词超出字数限制，请精简
高级检索

Characterization and substrate specificity study of the novel (R)-amine transaminase MagAT

  • 期刊论文

  • 发表日期：

    2025年11月22日

查看全文
  • Xiaole Yang,
  • Xia Tian,
  • Hai Zhu,
  • Weiwei Jia,
  • Jiahuan Li,
  • Ruizhou Tang,
  • Chuanzhi Zhang,
  • Yang Cao,
  • Tingting Li

摘要

Chiral amines, as essential chiral building blocks in drug synthesis, present a considerable challenge in biomanufacturing due to the requirement for highly efficient stereoselective synthesis. In this study, we successfully cloned and heterologously expressed a novel (R)-amine transaminase, MagAT, from Mycolicibacterium agri. Systematic analysis showed optimal activity at pH 7.0, with the highest reaction rate occurring within 30 min at 50 ℃. However, considering overall thermal stability, 40℃ was selected as the operating temperature for subsequent experiments. Furthermore, the enzyme retained nearly 100% catalytic activity in the presence of 10% methanol, DMSO, and chloroform. Kinetic analysis demonstrated that MagAT possessed high substrate affinity, with Michaelis constants (Km) of 5.13 ± 0.45 mM for (R)-1-phenylethan-1-amine and 3.73 ± 0.34 mM for pyruvate. Substrate specificity studies indicated that MagAT efficiently catalyzed the conversion of C3-C8 aliphatic amines (e.g., (R)-2-aminooctane) and monocyclic aromatic amines (e.g., (R)-1-phenylethan-1-amine), whereas its catalytic activity significantly declined for sterically hindered substrates such as naphthylamine. 3D structural modeling with AlphaFold3 and molecular docking showed that steric hindrance from key residues in the O-pocket critically influences substrate specificity. This study not only provides an efficient biocatalyst for the biosynthesis of chiral amines but also establishes a theoretical basis for the rational design of transaminases with broader substrate adaptability.