(593s) Deconstruction of Insulin Amyloid Fibrils Upon Laser Irradiation and Small Molecules and Probed by Time-Resolved Fluorescence Spectroscopy

The accumulation of misfolded proteins as amyloid fibrils is the hallmark of a large number of human degenerative diseases [Gotz, 2009]. Recent evidence indicates that protein can induce photophysical changes to the amyloid indicator dye thioflavin T, indicating that the active sites of different amyloid proteins may be the same [Mishra, 2008; Arora et al, 2004]. The photoinhibition mechanism of different ThT analogues on amyloid proteins is therefore of great importance. Here, we targeted insulin fibril formation by small aromatic molecules, with different numbers of aromatic rings and various substituents, and studied the requisite probes for the identification of amyloid fibrils and the natural plant polyphenols for inhibition of protein aggregation in view of their aromatic structures.

The interactions between small aromatic molecules and insulin amyloid fibrils were monitered by Florescence Resonance Energy Transition (FRET) method. Steady-state fluorescence results indicate that insulin amyloid fibrils are acceptors and tyrosine is the active site, transferring energy to the around donors. The inhibition effects of small aromatic molecules on insulin fibrils were observed by laser irradiation, which were further confirmed by the time-resolved fluorescence studies. The docking results further indicated that congo red, ThT, and resveratrol showed high inhibitory effects on protein aggregation when conjugated with laser irradiation, whereas procyanidin and trehalose could not disrupt insulin amyloid fibrils. Our FRET results may provide an effective way to design potential inhibitors of amyloid fibrils.

This work was supported by the Natural Science Foundation of China (No. 20806057 and 31071509), the Ministry of Science and Technology of China (Nos. 2012BAD29B05 and 2012AA06A303), and the Ministry of Education (No. NCET-11-0372).

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