【作者】 刘承智； 谢湖均； 许勇泉；

【Author】 Liu Chengzhi;Xie Hujun;Xu Yongquan;School of Food Science and Biotechnology,Zhejiang Gongshang University;Tea Research Institute,Chinese Academy of Agricultural Sciences;

【机构】 浙江工商大学食品与生物工程学院； 中国农业科学院茶叶研究所；

【摘要】 pH值对β-LG的聚集状态有重要的影响,利用多光谱学和分子动力学模拟方法研究了pH调控的β-LG与EGCG的相互作用机制。UV-Vis中观察到吸光度的增加和最大波长的蓝移,证实了β-LG-EGCG复合物的形成。荧光数据发现EGCG对β-LG的猝灭主要为静态猝灭,结合亲和力顺序为pH 7(K_a=1.83×10~4)> pH 5.3(K_a=1.6×10~4)>pH 2.5(K_a=1.10×10~4),这与分子动力学模拟得到的结果相一致。FTIR和圆二色谱(CD)研究表明,EGCG与β-LG之间的相互作用使得β-LG的二级结构发生了轻微的变化。分子动力学模拟结果表明,在pH值为7(二聚体)和5.3(四聚体)时,EGCG的首选结合位点位于由两个I片层和α螺旋组成的结合口袋中。而在pH值为2.5(单体)时,由于β-LG封闭的β-桶状结构,EGCG的结合位点位于β-LG的外表面。在不同的pH值下(2.5、5.3和7),β-LG-EGCG复合物的SASA值均低于β-LG,表明β-LG与EGCG结合后相对稳定,因为与单独的β-LG相比,结合有EGCG分子的β-LG分子内部埋藏了更多的疏水残基。导致EGCG与β-LG疏水区域的结合更紧密,SASA值更小,这与荧光数据相吻合。更多还原

【Abstract】 pH values have an important influence on the aggregation state of β-lactoglobulin(β-LG).In this study,the pH-dependent interaction mechanisms between β-LG and epigallocatechin-3-gallate(EGCG) were explored by multispectroscopy and molecular dynamics simulation.Fluorescence data showed that the quenching of β-LG by EGCG was mainly static quenching at different pH values,and the interaction between β-LG and EGCG was endothermic and spontaneously driven by hydrophobic interactions.Molecular dynamics(MD) simulation elucidated that EGCG preferred to bind to the pocket of β-LG at pH 7.0(dimer) and 5.3(tetramer),which consisted of two I lamellae and an α-helix.However,the binding site of EGCG at pH 2.5(monomer) was located on the outer surface of β-LG due to the closure of β-barrel structure ofβ-LG.Moreover,the binding free energies had a trend of pH 7.0> pH 5.3> pH 2.5,which was consistent with the trend of thermodynamic data.This study revealed the interaction mechanisms between EGCG and β-LG,which is of great significance for further development of dairy products as EGCG delivery system.更多还原