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Shedding light on electrostatic interactions of thioflavin T (ThT) and amyloids

Since the nineteenth century the benzothiazole-based small molecular rotor thioflavin T (ThT) has been the gold standard indicator for amyloidogenic assemblies¹. Amyloids that are formed due to protein misfolding into fibrillar aggregates mostly in β-sheet structure, are found in various diseases such as Alzheimer’s, Parkinson`s, type II diabetes, Amyloidosis and other neurodegenerative diseases. While the fluorescence increase of ThT upon binding to β-sheet structures is well established, its affinity to different proteins and amyloids remains unclear^2,3.

Herein we report the high impact of charge on binding ability of ThT to β-sheet structure. We use amphiphilic and charged β-sheet model peptides to obtain fibril`s with different surface charge states and monitored their interactions with ThT. Likewise, we demonstrate the influence of charged state on ThT binding by different β-amyloid fibrils. Using advanced spectroscopic techniques, combined with electron microscopy and fluorescent imaging, the structure of the amyloid-like assemblies was monitored in solution as function of pH and ζ potential measurements.

Results demonstrate the dramatic effect charged surface may have on ThT fluorescence intensity that should be taken into consideration in ThT quantitative measurement of amyloids.

References:

(1) Wolfe, L. S., Calabrese, M. F., Nath, A., Blaho, D. V, Miranker, A. D., and Xiong, Y. (2010) Protein-induced photophysical changes to the amyloid indicator dye thioflavin T. Proc. Natl. Acad. Sci. 107, 16863–16868.

(2) Amdursky, N., Erez, Y., and Huppert, D. (2012) Molecular rotors: What lies behind the high sensitivity of the thioflavin-T fluorescent marker. Acc. Chem. Res. 45, 1548–1557.

(3) Young, L. M., Ashcroft, A. E., and Radford, S. E. (2017) Small molecule probes of protein aggregation. Curr. Opin. Chem. Biol. 39, 90–99.