Henan Provincial Engineering Research Center of Insect Bio-reactor, College of Life Science, Nanyang Normal University, 1638 Wolong Road, Nanyang, Henan 473061, People's Republic of China.
School of Bioengineering, Dalian University of Technology, 2 Linggong Road, Dalian, Liaoning 116024, People's Republic of China.
J Agric Food Chem. 2024 Oct 23;72(42):23333-23344. doi: 10.1021/acs.jafc.4c05072. Epub 2024 Oct 9.
Formate dehydrogenase can be utilized as a biocatalyst in the bioelectrocatalysis of converting CO into formic acid. However, its industrial application has been hindered by limited thermal stability. This study successfully obtained a mutant (D533S/E684I) with enhanced thermal stability and catalytic activity through the rational design of flexible regions. The mutant exhibited a half-life () 1.5 times longer than the wild type (WT) at 35 °C, along with a specific enzyme activity 7.46 times higher than that of the WT. Additionally, the catalytic efficiency (/ value) of the mutant toward the substrate was 2.72 s·mM, representing a 19.4-fold increase compared to the WT (0.14 s·mM). Formic acid production reached 53.4 mM through bioelectrocatalysis after 10 h, utilizing the mutant as the biocatalyst. Molecular dynamics simulations and structural analysis were employed to investigate the molecular mechanisms behind the enhanced thermal stability and activity. The displacement of a highly flexible region in the mutant may counteract the stability-activity trade-off. This study proposed a method for improving both thermal stability and activity in enzyme evolution.
甲酸脱氢酶可以作为一种生物催化剂，用于将 CO 生物电化学转化为甲酸。然而，其工业应用受到热稳定性有限的限制。本研究通过对柔性区域的合理设计，成功获得了一种热稳定性和催化活性增强的突变体（D533S/E684I）。与野生型（WT）相比，该突变体在 35°C 下的半衰期（）延长了 1.5 倍，比 WT 的比酶活高 7.46 倍。此外，该突变体对底物的催化效率（/ 值）为 2.72 s·mM，比 WT（0.14 s·mM）提高了 19.4 倍。通过生物电化学催化，该突变体作为生物催化剂，10 小时后可生产 53.4 mM 甲酸。采用分子动力学模拟和结构分析研究了增强热稳定性和活性的分子机制。突变体中一个高度柔性区域的位移可能抵消了稳定性-活性权衡。本研究提出了一种在酶进化中同时提高热稳定性和活性的方法。
