Department of Biochemical Engineering, School of Chemical Engineering and Technology and, Key Laboratory of Systems Bioengineering and, Frontiers Science Center for Synthetic Biology (Ministry of Education), Tianjin University, Tianjin, 300350, China.
Chembiochem. 2023 Nov 2;24(21):e202300373. doi: 10.1002/cbic.202300373. Epub 2023 Sep 13.
Polyethylene terephthalate (PET) is one of the most widely used plastics, and the accumulation of PET poses a great threat to the environment. IsPETase can degrade PET rapidly at moderate temperatures, but its application is greatly limited by the low stability. Herein, molecular dynamics (MD) simulations combined with a sequence alignment strategy were adopted to introduce salt bridges into the flexible region of IsPETase to improve its thermal stability. In the designed variants, the T values of IsPETase and IsPETase were 7.4 and 8.7 °C higher than that of the wild type, respectively. The release of products degraded by IsPETase was 4.3 times that of the wild type. Tertiary structure characterization demonstrated that the structure of the variants IsPETase and IsPETase became more compact. Extensive MD simulations verified that a stable salt bridge was formed between the residue R168 and D186 in IsPETase , while in IsPETase an R168-D186-E188 salt bridge network was observed. These results confirmed that the proposed computation-based salt bridge design strategy could efficiently generate variants with enhanced thermal stability for the long-term degradation of PET, which would be helpful for the design of enzymes with improved stability.
聚对苯二甲酸乙二醇酯（PET）是应用最广泛的塑料之一，PET 的积累对环境构成了巨大威胁。对苯二甲酸酯酶（PETase）可以在中等温度下快速降解 PET，但由于其稳定性低，其应用受到极大限制。在此，采用分子动力学（MD）模拟结合序列比对策略，在 IsPETase 的柔性区域引入盐桥，以提高其热稳定性。在设计的变体中，IsPETase 和 IsPETase 的 T 值分别比野生型高 7.4 和 8.7°C。IsPETase 降解产物的释放量是野生型的 4.3 倍。三级结构特征表明，变体 IsPETase 和 IsPETase 的结构变得更加紧凑。广泛的 MD 模拟证实，在 IsPETase 中，残基 R168 和 D186 之间形成了一个稳定的盐桥，而在 IsPETase 中观察到 R168-D186-E188 盐桥网络。这些结果证实，所提出的基于计算的盐桥设计策略可以有效地产生具有增强热稳定性的变体，用于 PET 的长期降解，这将有助于设计具有改进稳定性的酶。
