Study on the binding of Thioflavin T to β-sheet-rich and non-β-sheet cavities

Amyloid fibril formation plays a role in more than 20 diseases including Alzheimer’s disease. In vitro detection of these fibrils is often performed using Thioflavin T (ThT), though the ThT binding mode is largely unknown. In the present study, spectral properties of ThT in binding environments representing β-sheet-rich and non-β-sheet cavities were examined. Acetylcholinesterase and γ-cyclodextrin induced a characteristic ThT fluorescence similar to that with amyloid fibrils, whereas β-cyclodextrin and the β-sheet-rich transthyretin did not. The cavities of acetylcholinesterase and γ-cyclodextrin were of similar diameter and only these cavities could accommodate two ThT ions according to molecular modelling. Binding stoichiometry studies also showed a possible binding of two ThT ions. Thus, the characteristic ThT fluorescence is induced in cavities with a diameter of 8–9 Å and a length able to accommodate the entire length of the ThT ion. The importance of a cavity diameter capable of binding two ThT ions, among others, indicates that an excimer formation is a plausible mechanism for the characteristic fluorescence. We propose a similar ThT binding mode in amyloid fibrils, where cavities of an appropriate size running parallel to the fibril axis have previously been proposed in several amyloid fibril models.