Novel method for quantitative determination of amyloid fibrils of alpha-synuclein and amyloid beta/A4 protein by using resveratrol.
Amyloidosis producing insoluble fibrillar protein aggregates is the common pathological feature of various neurodegenerative disorders such as Parkinson's and Alzheimer's diseases in which alpha-synuclein and amyloid beta/A4 protein (Abeta) participate to form Lewy bodies and senile plaques, respectively. To develop a novel analytical tool for amyloidosis, resveratrol, the major phenolic constituent of red wine and isolatable from grapevines, was employed to monitor the amyloids of alpha-synuclein and Abeta. Specific interaction to the amyloids enhanced the intrinsic fluorescence of resveratrol at 395 nm with an advent of new shoulder peak at 440 nm following an excitation at 320 nm. An increase in the resveratrol binding fluorescence was proportional to the quantity of amyloids. Typical sigmoidal kinetics of the amyloidosis of alpha-synuclein assessed with the thioflavin-T binding fluorescence or the beta-sheet content was fully reproduced by the resveratrol binding fluorescence. The resveratrol binding to the amyloids became saturated as the dye concentration increased, whereas the enhanced thioflavin-T binding fluorescence was quenched by the unbound thioflavin-T at the high dye concentration. Because resveratrol does not require any adjustment of the amyloid/dye ratio to obtain optimal amyloid binding fluorescence, and it exerts a higher quantum yield than does thioflavin-T, resveratrol is suggested to be a specific and more reliable fluorescent probe to determine the amyloids quantitatively.
Ahn
Jung Sun
JS
School of Chemical and Biological Engineering, College of Engineering, Seoul National University, Kwanak-Ku, Seoul 151-744, Korea.
Lee
Jung-Ho
JH
Kim
Je-Hoon
JH
Paik
Seung R
SR
D000682:Amyloid / Q000032:analysis*; D016229:Amyloid beta-Peptides / Q000032:analysis*; D000686:Amyloidosis / Q000175:diagnosis; D052160:Benzothiazoles; D006801:Humans; D020836:Protein Structure, Quaternary; D000077185:Resveratrol; D013050:Spectrometry, Fluorescence; D013267:Stilbenes / Q000737:chemistry*; D013844:Thiazoles / Q000737:chemistry; D051844:alpha-Synuclein / Q000032:analysis*
D000682:Amyloid; D016229:Amyloid beta-Peptides; D052160:Benzothiazoles; D013267:Stilbenes; D013844:Thiazoles; D051844:alpha-Synuclein; C009462:thioflavin T; D000077185:Resveratrol