Mind Your Dye: The Amyloid Sensor Thioflavin T Interacts with Sulfated Glycosaminoglycans Used To Induce Cross-β-Sheet Motifs

Abstract

Thioflavin T (ThT) is a commonly employed fluorescence probe for sensing amyloid fibrils. These highly ordered, insoluble nanostructures colocalize with sulfated glycosaminoglycans (GAGs) being abundant in the extracellular matrix and on the outer surface of cell membranes. To elucidate the positive impact of GAGs on amyloidogenesis, they are frequently used to promote fibril formation in vitro, which is detected by the enhanced fluorescence of ThT. The polyanionic nature and the affinity of GAGs to basic compounds predict that cationic ThT molecules may also bind to them, in addition to cross-β-structures formed in the reaction medium. By means of circular dichroism (CD) and absorption spectroscopy, this study examined the heparin and chondroitin sulfate binding of ThT. The large blue shift of the absorption peak indicated a card-pack type oligomerization of the dye molecules along the linear GAG chains. The strong exciton couplet observed in the CD spectra implies the left-handed, helical arrangement of GAG-associated oligomers of the dye. The decisive contribution of ionic forces for the binding was illustrated by sodium-ion-provoked dissociation of dye−GAG complexes. In silico analysis was performed to complement the experimental findings and to contribute to the understanding of potential molecular mechanisms underlying ThT−GAG interactions. ThT can be considered as an inert component in GAG-induced amyloid assays but only if the experiments are correctly designed.

Exploring the Nucleobase-Specific Hydrophobic Interaction of Cryptolepine Hydrate with RNA and Its Subsequent Sequestration

“…Figure B shows a strong negative peak at ∼282 nm, and the ellipticity of the strong negative signal increases with increasing RNA concentration. This observation leads to the conclusion that the orientation of the intercalated drug molecules within the RNA scaffolds is such that the long axis of the drug molecule is parallel to the long axis of the RNA base pair (a positive peak in the induced CD profile would have indicated a perpendicular orientation of the long axis of the drug molecule with respect to the long axis of RNA base pair). , The induced negative (at ∼282/378 nm) and positive (at ∼302 nm) peaks of CRYP in the presence of RNA may be a signature of the intermolecular exciton coupling after its interaction with RNA emerging from the close proximity of the CRYP molecules within the RNA-bound state. , The intermolecular exciton of CRYP may interact through space after its interaction with RNA and thus grows chirality along the axis of the RNA structure. , …”

Comment on “Emissive H-Aggregates of an Ultrafast Molecular Rotor: A Promising Platform for Sensing Heparin”

“…Additionally, the bell-shaped absorption band of the heparin-free dye at 413 nm exhibited a large blue shift to 393 nm and concomitant intensity reduction (hypochromism) . The absorption spectral changes are characteristic of dipole–dipole exciton coupling between the electronic transition moments of adjacent ThT molecules forming card-pack-type assemblies upon heparin binding . As a benzothiazole-type cationic dye, ThT bears a positively charged, quaternary nitrogen atom that is attracted electrostatically by the densely sulfated disaccharide units of heparin (two or three sulfate groups per repeat unit).…”

“…1 The absorption spectral changes are characteristic of dipole−dipole exciton coupling between the electronic transition moments of adjacent ThT molecules forming card-pack-type assemblies upon heparin binding. 2 As a benzothiazole-type cationic dye, ThT bears a positively charged, quaternary nitrogen atom that is attracted electrostatically by the densely sulfated disaccharide units of heparin (two or three sulfate groups per repeat unit). The decisive contribution of intermolecular ionic forces in the stabilization of ThT-heparin complexes was clearly demonstrated by the authors.…”

“…The ionic strength dependence of the ThT-heparin interaction is in full agreement with our recent circular dichroism (CD) spectroscopic studies. 2 Induced CD signals of the heparin associated dye showed high sensitivity to sodium ion concentration of the sample solution. The magnitudes of the ellipticity bands steadily decreased in parallel with the salt addition and canceled completely at ∼3 mM NaCl.…”

“…The ionic strength dependence of the ThT-heparin interaction is in full agreement with our recent circular dichroism (CD) spectroscopic studies . Induced CD signals of the heparin associated dye showed high sensitivity to sodium ion concentration of the sample solution.…”