A Label-Free Fluorometric Glutathione Assay Based on a Conformational Switch of G-quadruplex.

Xi Zhou,Doudou Zhang,Changbei Ma,Hailun He,Ying Yan,Yukui Zhou

doi:10.3390/molecules26092743

Abstract

In this paper, a label-free fluorescent method for glutathione (GSH) detection based on a thioflavin T/G-quadruplex conformational switch is developed. The sensing assay is fabricated depending on the virtue of mercury ions to form a thymine–thymine mismatch, which collapses the distance between two ssDNA and directs the guanine-rich part to form an intra-strand asymmetric split G-quadruplex. The newly formed G-quadruplex efficiently reacts with thioflavin T and enhances the fluorescent intensity. In the presence of GSH, Hg2+ is absorbed, destroying the G-quadruplex formation with a significant decrease in fluorescence emission. The proposed fluorescent assay exhibits a linear range between 0.03–5 μM of GSH with a detection limit of 9.8 nM. Furthermore, the efficacy of this method is examined using human serum samples to detect GSH. Besides GSH, other amino acids are also investigated in standard samples, which display satisfactory sensitivity and selectivity. Above all, we develop a method with features including potentiality, facility, sensitivity, and selectivity for analyzing GSH for clinical diagnostics.

Highlights

Glutathione (GSH), called γ-glutamyl-cysteinyl-glycine, is a thiol tripeptide containing glutamic acid, cysteine, and glycine [1]
It is well known that the G-quadruplex is formed by G-rich sequences, and it significantly increases the fluorescence intensity when reacting with small molecules, including thioflavin T (ThT), thiazole orange (TO), and N-methyl mesoporphyrin IX (NMM) [19,20,21,22]
According to the suggested method, Hg2+ interacts with a thymine–thymine mismatch to form T-Hg-T pairs to close the distance between DNA1 and DNA2 and directs two G-rich sides to construct a split

Summary

Introduction

Glutathione (GSH), called γ-glutamyl-cysteinyl-glycine, is a thiol tripeptide containing glutamic acid, cysteine, and glycine [1]. Glutathione plays critical roles in many cellular processes, such as cell growth and metabolism, and, notably, it governs the redox state of organs [3]. It is a kind of endogenous detoxicant; the active thiol can react with different components, especially heavy metals [1,4,5]. A fluorescent nanoprobe was proposed by Yang Liu to realize GSH, which is dependent on the coassembly of fluorescein and amphiphilic BODIPY [18] These assays mentioned above always need a complex process, or sensitivity is restrained, so an easier fluorescent strategy that has high sensitivity should be developed. It is well known that the G-quadruplex is formed by G-rich sequences, and it significantly increases the fluorescence intensity when reacting with small molecules, including thioflavin T (ThT), thiazole orange (TO), and N-methyl mesoporphyrin IX (NMM) [19,20,21,22]

Methods

Results

Conclusion