3种黄酮类化合物对胰脂肪酶的抑制作用及其结合机制

Inhibitory Effects of Three Flavonoids on Pancreatic Lipase and Their Binding Mechanism

探究小分子对胰脂肪酶(PL)的抑制作用已成为体外筛选具有抗肥胖作用活性成分的重要方法。采用分子对接法从黄酮类化合物中筛选具有强结合作用的活性成分,通过酶动力学研究抑制活性和抑制类型,结合荧光光谱法研究活性成分与PL的结合反应及机制。通过体外筛选得到3种黄酮类化学物:黄芩素(Bai)、槲皮素(Que)和漆黄素(Fis)。分子对接结果表明,3种黄酮类化合物与PL的结合位于催化位点Ser153-His264-Asp177残基附近,主要作用力为疏水相互作用和氢键。Bai、Que和Fis对PL抑制活性呈现剂量依赖性,其IC50分别为49.14,57.78,61.26μg/mL。抑制动力学表明:Bai和Fis对PL为竞争型抑制,而Que属于混合型抑制。荧光光谱法研究表明:3种黄酮类化合物对PL猝灭均属于静态猝灭,结合常数均大于105数量级,说明Bai、Que和Fis均与PL形成稳定的复合物。热力学参数分析表明:Bai与PL结合主要作用力为氢键和范德华力,Que、Fis和PL结合主要通过疏水相互作用。本研究将为开发新型PL抑制剂提供新思路。

The inhibition of small molecules on pancreatic lipase(PL)has become an important model for screening anti-obesity ingredients in vitro.In this paper,the active constituents with strong binding effect were screened from flavonoids by molecular docking method.The inhibitory activity and type of inhibition were studied by enzyme kinetics,and the binding reaction and mechanism of active constituents and PL were studied by fluorescence spectroscopy.Three flavonoids were selected in vitro:Baicalein(Bai),Quercetin(Que)and Fisetin(Fis).Molecular docking results showed that the binding of the three flavonoids to PL was located near the residues of Ser153-His264-Asp177 at the catalytic site,and the main forces were hydrophobic interaction and hydrogen bonding.Bai,Que and Fis showed dose-dependent inhibitory activity on PL,with IC50 values of 49.14,57.78,61.26μg/mL,respectively.Inhibition kinetics showed that Bai and Fis had competitive inhibition on PL,while Que was mixed inhibition.Fluorescence spectroscopy showed that the quenching of PL by the three flavonoids belonged to static quenching,and the binding constants were all greater than 105 orders of magnitude,indicating that Bai,Que and Fis formed stable complexes with PL.The analysis of thermodynamic parameters showed that the main binding forces of Bai and PL were hydrogen bond and van der Waals force,and the binding of Que,Fis and PL was through hydrophobic interaction.This paper will provide new ideas for the development of novel PL inhibitors.