Molecular Interactions Between Amyloid Proteins And (–)-Epigallocatechin-3-Gallate

The conversion of amyloid proteins from native forms to oligomers and fibrils isthe hallmark of many neurodegenerative diseases.(–)-Epigallocatechin-3-gallate(EGCG) has been reported to effectively prevent the onset of many neurodegenerativediseases. However, the interactions between EGCG and amyloid proteins are stillunclear. Herein, isothermal titration calorimetry (ITC), circular dichroism, massspectroscopy, transmission electron microscopy, high performance liquidchromatography, etc were used to investigate the molecular interactions betweenEGCG and two amyloid proteins, i.e., amyloid β-protein (Aβ42) and human insulin(hIns) to facilitate the rational screening and designing of amyloid aggregationinhibitors.The thermodynamic interactions between EGCG and Aβ42as well as its threefragments, i.e., Aβ1-16, Aβ1-30, and Aβ31-42, were studied by ITC. The results showthat hydrogen bonding and hydrophobic interaction are both substantial in the binding.Hydrogen bonding mainly happens in Aβ1-16while hydrogen bonding mainlyhappens in Aβ17-42. With the increase of EGCG concentration, the relativecontribution of hydrogen bonding decreases while that of hydrophobic interactionincreases. The binding can be promoted by increasing temperature at about100mmol/L NaCl and pH values away from Aβ42’s isoelectric point.Hydrogen bonding is always involved in the binding of EGCG to hIns. However,with the increase of EGCG concentration, the contribution of hydrophobic interactionincreases. There is a large EEC in the interactions between EGCG and hIns. EGCGpreferentially binds to hIns, decreasing the hydrodynamic radius of hIns and slightlychanging the secondary structure of hInsThe inhibitory effects of EGCG on the fibrillation of hIns at pH2.0and60°Cfirstly increase with increasing EGCG concentration and then become constant.EGCG reduces the length and width of fibrils. At pH7.4and37°C, EGCG redirectshIns into globular aggregates, and the inhibitory effect reaches maximum at0.1-0.2mmol/L EGCG. EGCG keeps part of hIns from aggregation and slows down thechanges of the secondary structures of hIns in the aggregation. Finally, two physicalmodels were proposed to explain the molecular interactions between hIns and EGCGat the two conditions.1-5mM EGCG induced hIns into reversible globular precipitates of185to365nm. The results show that hydrophobic interaction was the main driving force andEGCG clusters play an important role in inducing hIns precipitation. The precipitatecan be completely dissolved by0.67to5.8M urea and the secondary structure of hInscan also recover after52-h incubation. Based on extensive investigation, a physicalmodel was proposed to explain the molecular interactions between EGCG and hIns.