多光谱法结合分子对接探究4种黄酮与β-乳球蛋白的相互作用

Exploration on interaction between 4 kinds of flavonoids and β-lactoglobulin by multi-spectroscopy analysis and molecular docking

目的 探究4种黄酮[柚皮苷(naringin,Nar)、黄芩素(baicalein,Bai)、白杨素(chrysin,Chr)和漆黄素(fisetin,Fis)]与β-乳球蛋白(β-lactoglobulin,β-Lg)的相互作用。方法 首先采用分子对接技术模拟预测4种黄酮与β-Lg的结合位点,再利用荧光光谱法研究4种黄酮与β-Lg的荧光猝灭作用及作用机制,并计算相关参数,最后通过多种光谱法分析4种黄酮对β-Lg空间构象的变化。结果 分子对接结果 表明Nar和Fis结合在β-Lg的疏水空腔口处,Bai与Chr与β-Lg的结合位点在β-Lg的外表面。荧光光谱结果 表明,4种黄酮对β-Lg均产生静态猝灭,结合常数均在104~107数量级,说明Nar/Bai/Chr/Fis均与β-Lg结合紧密且形成稳定的复合物;热力学参数分析表明Nar/Bai/Chr/Fis与β-Lg的结合为自发进行,其中Nar/Bai与β-Lg作用力类型是氢键和范德华力, Chr/Fis与β-Lg是疏水相互作用。同步荧光光谱、三维荧光光谱、紫外-可见光吸收光谱和傅里叶变换红外光谱结果 表明,Nar/Bai/Chr/Fis与β-Lg发生相互作用后使β-Lg的空间构象发生变化。结论 4种黄酮均能与β-Lg结合,为黄酮类化合物与生物大分子作用机制研究及对活性成分的保护和利用提供理论支持。

Objective To explore the interaction between 4 kinds of flavonoids,namely naringin(Nar),baicalein(Bai),chrysin(Chr),fisetin(Fis)andβ-lactoglobulin(β-Lg).Methods In this study,the binding sites of 4 different types of flavonoids andβ-Lg were initially simulated and predicted using molecular docking technology.Then,the mechanism and fluorescence quenching effect of these 4 types of flavonoids andβ-Lg were investigated using fluorescence spectroscopy,and the pertinent parameters were calculated.Finally,the impact of 4 kinds of flavonoids on the changes in spatial conformation ofβ-Lg was analyzed using synchronous fluorescence spectroscopy,three-dimensional fluorescence spectrum,ultraviolet-visible absorption spectrum,and Fourier transform infrared spectroscopy.Results The results of molecular docking showed that Nar and Fis bind at the hydrophobic cavity opening ofβ-Lg and the binding site of Bai with Chr toβ-Lg was on the outer surface ofβ-Lg.The results of fluorescence spectrum indicated that all 4 flavonoids induced static quenching onβ-Lg,and the binding constants were around the range of 104-107,suggesting that Nar/Bai/Chr/Fis were firmly bound toβ-Lg and formed stable complexes.The investigation of thermodynamic parameters revealed that the binding betweenβ-Lg and Nar/Bai/Chr/Fis occurred spontaneously.Additionally,the interaction betweenβ-Lg and Nar/Bai involved hydrogen bonds and van der Waals forces,whileβ-Lg and Chr/Fis were bonded through hydrophobic interaction.The results of synchronous fluorescence spectroscopy,three-dimensional fluorescence spectrum,ultraviolet-visible absorption spectrum,and Fourier transform infrared spectroscopy demonstrated that the spatial conformation ofβ-Lg changed following the interaction betweenβ-Lg and Nar/Bai/Chr/Fis.Conclusion The findings demonstrate that each of the 4 flavonoids has the ability to bind toβ-Lg,offering theoretical backing for the investigation of the interaction between flavonoids and biological macromolecules as well as the preservation and application of active components.