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<p>This study used molecular dynamics simulations, B-factor analysis, and saturation mutagenesis screening to enhance the thermal stability of the <i>trans</i>-epoxysuccinate hydrolase (TESH) derived from <i>Pseudomonas koreensis</i>. Eleven mutants that influence this characteristic were selected, yielding four mutants with improved activity. Among them, mutants A142C and S178Q exhibited lower Michaelis constant (<i>K</i><sub><i>m</i></sub>) values, and their <i>k</i><sub>cat</sub>/<i>K</i><sub>m</sub> ratios (<i>k</i><sub>cat</sub>, catalytic constant) were 3.7 and 0.9 times higher than those of the wild type, respectively. The values of half-life at 50 °C <InlineEquation ID="IEq1"> <EquationSource Format="TEX">\((T_{1/2}^{50})\)</EquationSource> <EquationSource Format="MATHML"><math display="block"> <mo stretchy="false">(</mo> <msubsup> <mi>T</mi> <mrow> <mn>1</mn> <mrow> <mo>/</mo> </mrow> <mn>2</mn> </mrow> <mrow> <mn>50</mn> </mrow> </msubsup> <mo stretchy="false">)</mo> </math></EquationSource> </InlineEquation> of the two mutants were increased by 107% and 59%, respectively, compared to the wild type. Molecular docking and molecular dynamics simulations indicated that the two mutants showed stronger substrate interaction, lower binding energy, and reduced root mean square deviation compared to the wild type, along with decreased electrostatic potential energy and increased hydrophobicity near their mutation sites. The study of protein thermal stability engineering and associated mechanisms provides a valuable reference and holds practical significance for the industrial production of <i>meso</i>-tartaric acid.</p>

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Improving the thermal stability of trans-epoxysuccinate hydrolase

  • 期刊论文

  • 发表日期：

    2025年12月22日

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  • Wenna Bao,
  • Jinfeng Yao,
  • Haifeng Pan,
  • Ronglin Zhu,
  • Xinying Li,
  • Hongxiu Liao

摘要

This study used molecular dynamics simulations, B-factor analysis, and saturation mutagenesis screening to enhance the thermal stability of the trans-epoxysuccinate hydrolase (TESH) derived from Pseudomonas koreensis. Eleven mutants that influence this characteristic were selected, yielding four mutants with improved activity. Among them, mutants A142C and S178Q exhibited lower Michaelis constant (Km) values, and their kcat/Km ratios (kcat, catalytic constant) were 3.7 and 0.9 times higher than those of the wild type, respectively. The values of half-life at 50 °C \((T_{1/2}^{50})\) ( T 1 / 2 50 ) of the two mutants were increased by 107% and 59%, respectively, compared to the wild type. Molecular docking and molecular dynamics simulations indicated that the two mutants showed stronger substrate interaction, lower binding energy, and reduced root mean square deviation compared to the wild type, along with decreased electrostatic potential energy and increased hydrophobicity near their mutation sites. The study of protein thermal stability engineering and associated mechanisms provides a valuable reference and holds practical significance for the industrial production of meso-tartaric acid.