摘要
采用紫外-可见吸收光谱、荧光光谱和圆二色光谱并结合分子模拟技术,考察了槲皮素与胰蛋白酶(TRY)的作用机制及槲皮素的抗氧化活性。研究发现,槲皮素的加入可以猝灭TRY的内源性荧光,两者有一个结合位点,298K时结合常数为8.24×10~4 L·mol^(-1),且疏水作用力是驱动槲皮素与TRY结合的主要作用力。槲皮素能够诱导TRY的α-螺旋和无规则卷曲比率上升,而β-折叠和β-转角比率下降,导致二级结构发生变化。分子模拟结果显示,槲皮素分子的苯环结合到TRY疏水腔中,与Leu155,Pro28,Pro198,Val200,Ser 202和Cys157等主要氨基酸发生作用。抗氧化性实验表明,槲皮素对DPPH和羟基自由基的清除能力比维生素C强,而TRY对槲皮素清除DPPH自由基的能力产生抑制作用。
The interaction between quercetin and trypsin was studied by UV-vis absorption spectroscopy,fluorescence spectroscopy and circular dichroism（CD）spectroscopy coupled with molecular simulation.It was found that quercetin has a strong fluorescence quenching effect on trypsin,and the fluorescence quenching was a static mechanism.There was a single class of quercetin binding sites on trypsin,and the binding constant was 8.24×10~4 L·mol-1 at 298 K,indicating that a moderate affinity existed between quercetin and TRY.The calculated thermodynamic parameters at different temperatures suggested that hydrophobic interaction was the main driving force in the binding process.The UV-vis absorption and CD spectra indicated that the addition of quercetin resulted in the conformational alteration of trypsin with an increase inα-helix and random coil,and a decrease inβ-sheet andβ-turn structure.The molecular simulation results showed that the benzene ring of the quercetin bound to the hydrophobic cavity of trypsin,and interacted with the main amino acid residues such as Leu155,Pro28,Pro198,Val200,Ser 202 and Cys157.The results of antioxidant experiments showed that quercetin had strong DPPH and hydroxyl radical scavenging ability,and its ability to scavenge the two kinds of free radical was stronger than that of positive control vitamin C.Trypsin was found to inhibit the DPPH radical scavenging ability of quercetin.
出处
《南昌大学学报（理科版）》
CAS
北大核心
2017年第3期244-248,254,共6页
Journal of Nanchang University(Natural Science)
基金
国家自然科学基金项目(31460422
31060210)
江西省自然科学基金项目(20171BAB204029
20143ACB20006)
江西省教育厅科技项目(GJJ150187)
食品科学与技术国家重点实验室目标导向课题(SKLF-ZZA-201612)
关键词
槲皮素
胰蛋白酶
荧光猝灭
分子模拟
自由基清除
quercetin
trypsin
fluorescence quenching
molecular simulation
free radicals
