Effect of (−)-epigallocatechin-3-gallate on human insulin fibrillation/aggregation kinetics

Abstract

(−)-Epigallocatechin-3-gallate (EGCG), a food additive derived from green tea, has been reported to effectively inhibit the fibrillation of many amyloid proteins, but not insulin. So herein, the influences of EGCG on the fibrillation kinetics of human insulin at two conditions (pH 2.0, 60°C and pH 7.4, 37°C) were extensively studied. It was found that at pH 2.0 and 60°C the inhibitory effect increased with increasing EGCG concentration from 0.35 to 3.5mmol/L but kept almost unchanged from 3.5 to 5mmol/L EGCG. The addition of EGCG reduced the length and width of fibrils and kept part of insulin from fibrillation at this condition. At pH 7.4 and 37°C, however, EGCG altered the fibrillation pathway of insulin and redirected it into globular aggregates, and the inhibitory effect of EGCG on the aggregation reached maximum at about 0.1–0.2mmol/L. In this case, part of insulin molecules were prevented from aggregation and existed as a mixture of monomer, dimer, tetramer, and hexamer in the solution. Circular dichroism spectroscopy indicated that EGCG slowed down the changes of the secondary structures of insulin in the aggregation. Finally, two physical models were proposed to explain the molecular interactions between insulin and EGCG at the two conditions. The research has clarified the kinetic mechanism of the inhibitory effect of EGCG on insulin fibrillation/aggregation.