Minimal Thioflavin T Modifications Improve Visual Discrimination of Guanine-Quadruplex Topologies and Alter Compound-Induced Topological Structures

Abstract

We newly synthesized thioflavin T (ThT) analogs for which the methyl group at the N3 position on the benzothiazole ring was replaced with either a ((p-(dimethylamino)benzoyl)oxy)ethyl group (ThT-DB) or a hydroxyethyl group (ThT-HE). In several neutral buffers, ThT-HE bound to a parallel guanine-quadruplex (G4) DNA and selectively emitted strong fluorescence at 74- to 240-fold higher intensities than those in the presence of double-stranded DNA (dsDNA), whereas ThT resulted in only 13- to 25-fold higher intensities. Furthermore, circular dichroism (CD) analyses using ThT, ThT-DB, and ThT-HE showed that these compounds could induce topological changes in G4. In addition, the different chemical structures of the N3 substituents could alter a G4-DNA conformation. These results indicate a great potential for N3-substituted ThT analogs as G4 probes and drug leads to achieve gene expression regulation.

Fluorescent dyes in the context of DNA‐binding: The case of Thioflavin T

“…In that case, the photoexcited state cannot relax into the twisted one and a fluorescence light‐up is observed. Due to this characteristic, ThT has become a “gold standard” for visualizing and quantifying the presence of amyloid fibrils and it has been successfully used as a G‐quadruplex specific fluorescent sensor . Recent studies have also investigated the affinity of ThT binding to single and double stranded DNA …”

Thioflavin T as an Efficient G-Quadruplex Inducer for the Highly Sensitive Detection of Thrombin Using a New Föster Resonance Energy Transfer System

“…59 At present, the G-quadruplex specific ThT has attracted increasing interests, and it has been used as a light-up fluorescence probe for the sensing of the G-quadruplex, biothiols, Hg 2+ , K + , liver-related short gene, and the structure change of i-moif. [60][61][62][63][64][65] Inspired by these studies, we constructed a new FRET system using CP as an donor and ThT as an acceptor, and designed a simple, label-free, and sensitive biosensor for the detection of thrombin in buffer and in diluted serum. An improved sensitivity for thrombin detection was obtained when compared with two recent strategies based on the FRET from the CP to the dye.…”

Conjugating a groove-binding motif to an Ir(iii) complex for the enhancement of G-quadruplex probe behavior

“…S1a†). This is comparable to organic G-quadruplex probes, which exhibited a high fold-enrichment for G-quadruplex DNA and moderate enhancement for ssDNA and dsDNA23,41 Meanwhile, modified ThT analogues have been reported by Kuwahara's group with high G-quadruplex/ssDNA or G-quadruplex/dsDNA enhancement ratios 24…”

“…This approach has the disadvantage that fluorescently labeled oligonucleotides are relatively expensive, thus a more economical alternative may possibly involve a label-free strategy that utilizes unmodified DNA and selective DNA probes. Several organic dyes have been reported as DNA probes, such as thioflavin T (ThT),23,24 crystal violet (CV)25 and thiazole orange (TO),26 some of which can recognize G-quadruplex (G4) DNA via the end stacking of a G-quartet.…”