New Fluorescent Probe for Continuous Monitoring of Alpha-Synuclein Aggregation

Abstract

The aggregation of the presynaptic protein alpha-synuclein (AS), associated with Parkinson's disease, results in fibrils with a cross-beta-amyloid structure. Thioflavin T (ThT) is widely used for the identification and quantification of such amyloid fibrils in vitro. However, it exhibits poor sensitivity and reproducibility, requires sampling, and is insensitive to the early stages of aggregation.View Large Image | View Hi-Res Image | Download PowerPoint SlideWe introduced a new sensor molecule for the continuous monitoring of AS aggregation, denoted AS-140HC, consisting of the AS mutant (A140C, C-terminus) labeled with the 3-hydroxychromone dye MFC[1]. MFC exhibits two fluorescence bands (N∗ and T∗) arising from Excited State Intramolecular Proton Transfer (ESIPT)[2]. The intensity ratio of (IT∗/IN∗) reflects the microenvironment of the probe. Addition of AS-140HC in the range of 0.5-5% to wild type AS allows the monitoring of aggregation via the strong increase of IT∗/IN∗ (panel A), which occurs at a much earlier stage of aggregation than the ThT response (panel B). See also refs [3-6].[1] manuscript in preparation; [2] Demchenko et al., Biophys J., 2009, 3461; [3] poster Fauerbach et al.; [4] poster Shvadchak et al.; [5] Caarls et al., J. Fluor, 2009, DOI 10.1007/s10895-009-0536-1; [6] Celej et al., Biochemistry 2009, 7465.