Centre for Nutrition and Food Sciences, ARC Centre of Excellence in Plant Cell Walls, Queensland Alliance for Agriculture and Food Innovation, The University of Queensland, Queensland, Australia.
Quadram Institute, Norwich Research Park, Norwich, UK.
Mol Nutr Food Res. 2017 Oct;61(10). doi: 10.1002/mnfr.201700324. Epub 2017 Aug 23.
This study aims to use a combination of biochemical and biophysical methods to derive greater mechanistic understanding of the interactions between tea polyphenols and porcine pancreatic α-amylase (PPA).
The interaction mechanism was studied through fluorescence quenching (FQ), differential scanning calorimetry (DSC) and isothermal titration calorimetry (ITC) and compared with inhibition kinetics. The results showed that a higher quenching effect of polyphenols corresponded to a stronger inhibitory activity against PPA. The red-shift of maximum emission wavelength of PPA bound with some polyphenols indicated a potential structural unfolding of PPA. This was also suggested by the decreased thermostability of PPA with these polyphenols in DSC thermograms. Through thermodynamic binding analysis of ITC and inhibition kinetics, the equilibrium of competitive inhibition was shown to result from the binding of particularly galloylated polyphenols with specific sites on PPA. There were positive linear correlations between the reciprocal of competitive inhibition constant (1/K ), quenching constant (K ) and binding constant (K ).
The combination of inhibition kinetics, FQ, DSC and ITC can reasonably characterize the interactions between tea polyphenols and PPA. The galloyl moiety is an important group in catechins and theaflavins in terms of binding with and inhibiting the activity of PPA.
本研究旨在采用生化和生物物理方法相结合，深入了解茶多酚与猪胰α-淀粉酶（PPA）之间的相互作用机制。
通过荧光猝灭（FQ）、差示扫描量热法（DSC）和等温滴定量热法（ITC）研究了相互作用机制，并与抑制动力学进行了比较。结果表明，多酚的猝灭效应越高，对 PPA 的抑制活性越强。与一些多酚结合后 PPA 的最大发射波长红移表明 PPA 潜在的结构展开。这也可以从 DSC 图谱中这些多酚降低了 PPA 的热稳定性得到证实。通过 ITC 的热力学结合分析和抑制动力学，表明竞争性抑制的平衡是由于特定的没食子酰化多酚与 PPA 上的特定结合位点结合所致。竞争抑制常数的倒数（1/Ki）、猝灭常数（Kq）和结合常数（Kb）之间存在正线性相关关系。
抑制动力学、FQ、DSC 和 ITC 的组合可以合理地表征茶多酚与 PPA 之间的相互作用。没食子酰基是儿茶素和茶黄素中与 PPA 结合并抑制其活性的重要基团。
