光谱法、量热法以及分子对接研究酸枣仁皂苷A与中心蛋白的作用

Study on the interaction between centrin and jujuboside A by spectroscopic methodologies, calorimetric and molecular docking

中文关键词:  八肋游仆虫中心蛋白  酸枣仁皂苷A  圆二色光谱  三维荧光光谱  稳态荧光光谱  等温滴定量热法  分子对接技术

英文关键词:Euplotes octocarinatus centrin  jujuboside A  circular dichroism spectroscopy  three-dimensional fluorescence spectroscopy  steady-state fluorescence spectroscopy  isothermal titration calorimetry  molecular docking techniques

中文摘要:

      目的 研究酸枣仁皂苷A (jujuboside A, JuA)与八肋游仆虫中心蛋白(Euplotes octocarinatus centrin, EoCen)的相互作用。方法 采用圆二色光谱(circular dichroism spectroscopy, CD)以及三维荧光光谱研究EoCen的构象变化; 采用稳态荧光光谱与等温滴定量热法(isothermal titration calorimetry, ITC)研究JuA与EoCen作用的机制; 分子对接技术模拟JuA与EoCen的结合位点。结果 JuA以摩尔比1:1结合于EoCen的C端(二者反应的结合常数约为105 L/mol, 由ITC拟合结果可知, JuA与EoCen复合物的形成是放热的过程, 熵变对自由能的贡献较小, 主要由焓变驱动。EoCen与JuA结合后, 构象发生变化, α-螺旋含量减少, 酪氨酸残基的微环境变化, EoCen的C端不再结合蜂毒素。结论 JuA主要通过范德华力与EoCen结合, 从而改变EoCen的构象, 抑制其与靶蛋白的结合。

英文摘要:

      Objective To explore the interaction between jujuboside A (JuA) and Euplotes octocarinatus centrin (EoCen). Methods The conformational changes of EoCen were studied by circular dichroism spectroscopy (CD) and three-dimensional fluorescence spectroscopy. The interaction mechanism was studied by steady-state fluorescence spectroscopy and isothermal titration calorimetry (ITC). Molecular docking techniques simulated the binding sites of JuA and EoCen. Results Results indicated that JuA could form complex with the C-terminal domain of EoCen in molar ratio 1:1, and the conditional binding constant was 105 L/mol. The ITC fitting results showed that complex formation was an exothermic process, and the contribution of entropy changes to free energy was small, which was mainly driven by enthalpy change. Besides, the formation of EoCen-JuA complex resulted in a conformational change of protein. In more detail, the content of α-helix of protein decreased and microenvironmental of tyrosine residues changed. After binding to JuA, the C-terminal domain of EoCen could not bind to melittin. Conclusion JuA interacts with EoCen mainly by vander Waals force. Conformation of EoCen changes, thereby inhibiting the target peptide binding.