Submicron polymeric particles accelerate insulin fibrillation by surface adsorption.

Abstract

Insulin fibrillation is caused by the interaction of partially unfolded protein molecules resulting in the formation of insoluble aggregates. Engineered nano- and submicron particles have been reported to accelerate or inhibit the fibrillation of insulin. However, the mechanism by which engineered particles regulate insulin fibrillation kinetics is not fully understood. In the current study, insulin fibrillation in the presence of surface engineered, amine- or sulfate-modified polystyrene particles (200 nm) was investigated. Both particles were found to enhance the rate of protein fibrillation at high particle concentrations (1:1 mass ratio of proteins to particles) as evidenced by increased fluorescence of thioflavin T. A significant amount of proteins was found to adsorb on the surface of both particles, suggesting that the particles served as a nucleation site for fibrillation. Insulin fibrillation was associated with a change in the protein's secondary structure from α-helix to β-sheet. This change of structure was not significantly affected by the presence of particles. Monte Carlo simulations of protein fibrillation in the presence of different surfaces confirmed experimental results and demonstrated that protein fibrillation is enhanced as the affinity of protein molecules toward the surface increases. Along with increasing the local concentration of proteins, the presence of a surface stabilizes unfolded intermediates, which are precursors to fibrillation. These results provide insight into the mechanisms by which engineered particles promote protein fibrillation and can be important in the context of nanotoxicity.