State Key Laboratory of Food Science and Technology, Jiangnan University, Wuxi 214122, China; Department of Chemistry and Chemical Biology, Harvard University, Cambridge, MA 02138, United States.
State Key Laboratory of Food Science and Technology, Jiangnan University, Wuxi 214122, China.
Int J Biol Macromol. 2020 Jul 1;154:1356-1365. doi: 10.1016/j.ijbiomac.2019.11.015. Epub 2019 Nov 13.
Directed evolution using random mutation in vast sequence space leads to the low probability of obtaining target proteins. Emerging engineering strategies with computational tools are developed for more trustable outcomes. We used some semi-rational design methods to modify an industrial enzyme, namely cellobiose 2-epimerase (CE). A mutant was selected for its better thermostability and isomerization activity. The tradeoffs between thermostability, epimerization activity and isomerization activity of the CE mutants were different. To investigate the computational prediction performance of protein stability upon point mutations, molecular dynamics (MD) simulation analyses were conducted. The root mean square deviation (RMSD) and hydrogen bond analyses reproduced the correct trends in stability changes of the wild-type and mutated CEs with relatively high accuracy (correlation coefficients r ~ 0.5-0.8). The simulation temperature and time are important factors that influence the prediction performance. Our result shows that thermostability predictors calculated from MD simulation do better in predicting the thermostability changes of the mutated enzymes than the predictors using static-state information of the enzymes.
在广阔的序列空间中进行随机突变的定向进化导致获得目标蛋白的概率很低。新兴的工程策略与计算工具正在被开发出来，以获得更可靠的结果。我们使用了一些半理性设计方法来修饰一种工业酶，即纤维二糖 2-差向异构酶（CE）。选择了一个突变体，因为它具有更好的热稳定性和异构化活性。CE 突变体的热稳定性、异构化活性和异构化活性之间存在权衡。为了研究点突变后蛋白质稳定性的计算预测性能，进行了分子动力学（MD）模拟分析。均方根偏差（RMSD）和氢键分析以较高的准确性（相关系数 r~0.5-0.8）再现了野生型和突变型 CE 的稳定性变化的正确趋势。模拟温度和时间是影响预测性能的重要因素。我们的结果表明，与使用酶静态信息的预测因子相比，从 MD 模拟计算得出的热稳定性预测因子在预测突变酶的热稳定性变化方面表现更好。
