Identification of Thioflavin T Binding Modes to DNA: A Structure-Specific Molecular Probe for Lasing Applications.

P Hanczyc,B Feng,P Fita,P Rajchel-Mieldzioć

doi:10.1021/acs.jpclett.1c01254

P Hanczyc, B Feng + Show 2 more

Open Access

https://doi.org/10.1021/acs.jpclett.1c01254

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Abstract

The binding mechanism of thioflavin T (ThT) to DNA was studied using polarized light spectroscopy and fluorescence-based techniques in solutions and in solid films. Linear dichroism measurements showed that ThT binds to DNA duplex by intercalation. Time-resolved fluorescence studies revealed a second binding mode which is the external binding to the DNA phosphate groups. Both binding modes represent the nonspecific type of interactions. The studies were complemented with the analysis of short oligonucleotides having DNA cavities. The results indicate that the interplay between three binding modes—intercalation, external binding, and binding inside DNA cavities—determines the effective fluorescence quantum yield of the dye in the DNA structures. External binding was found to be responsible for fluorescence quenching because of energy transfer between intercalated and externally bound molecules. Finally, amplified spontaneous emission (ASE) was successfully generated in the ThT-stained films and used for detecting different DNA structures. ASE measurements show that ThT-stained DNA structures can be used for designing bioderived microlasers.

Highlights

The binding mechanism of thioflavin T (ThT) to DNA was studied using polarized light spectroscopy and fluorescence-based techniques in solutions and in solid films
It was found that ThT emission is highly sensitive to the local structure of surrounding biomolecules and their microenvironment.[5−7] That makes ThT a fluorophore widely used in biomedical research,[8] primarily to detect amyloid protein fibrils that are linked to neurodegenerative diseases.[9]
In this Letter, we examine the ThT interactions with four DNA samples: a natural long DNA duplex from calf thymus, and 28 bases long synthetic single- and double-stranded DNA, in which one of the single-stranded oligonucleotides mimics the classical DNA strand and the other, G-rich oligonucleotide, forms the G-quadruplex structure

Summary

Introduction

The binding mechanism of thioflavin T (ThT) to DNA was studied using polarized light spectroscopy and fluorescence-based techniques in solutions and in solid films. The emitting ThT species are located in DNA cavities and intercalation sites, whereas ThT molecules externally bound to the DNA phosphate groups quench the fluorescence of the former.

Results

Conclusion