DNA Binding Mode Analysis of a Core-Extended Naphthalene Diimide as a Conformation-Sensitive Fluorescent Probe of G-Quadruplex Structures.

Chiara Platella,Filippo Doria,Rosa Gaglione,Daniela Montesarchio,Valentina Pirota,Ettore Napolitano,Domenica Musumeci,Angela Arciello

doi:10.3390/ijms221910624

Abstract

G-quadruplex existence was proved in cells by using both antibodies and small molecule fluorescent probes. However, the G-quadruplex probes designed thus far are structure- but not conformation-specific. Recently, a core-extended naphthalene diimide (cex-NDI) was designed and found to provide fluorescent signals of markedly different intensities when bound to G-quadruplexes of different conformations or duplexes. Aiming at evaluating how the fluorescence behaviour of this compound is associated with specific binding modes to the different DNA targets, cex-NDI was here studied in its interaction with hybrid G-quadruplex, parallel G-quadruplex, and B-DNA duplex models by biophysical techniques, molecular docking, and biological assays. cex-NDI showed different binding modes associated with different amounts of stacking interactions with the three DNA targets. The preferential binding sites were the groove, outer quartet, or intercalative site of the hybrid G-quadruplex, parallel G-quadruplex, and B-DNA duplex, respectively. Interestingly, our data show that the fluorescence intensity of DNA-bound cex-NDI correlates with the amount of stacking interactions formed by the ligand with each DNA target, thus providing the rationale behind the conformation-sensitive properties of cex-NDI and supporting its use as a fluorescent probe of G-quadruplex structures. Notably, biological assays proved that cex-NDI mainly localizes in the G-quadruplex-rich nuclei of cancer cells.

Highlights

G-quadruplex structures are structural motifs of DNA or RNA, whose main unit involves a cyclic planar array of four guanines, which is called G-quartet [1,2,3,4]
As previously described [39], under the here used conditions, m-tel24 adopts a hybrid-1 G-quadruplex structural topology, involving G3:G21:G17:G9, G4:G10:G16:G22, and G5:G11:G15:G23 quartets, whereby G3, G9, G15, G16 and G21 are in syn conformation along with the N-glycosidic bond and the other residues are in anti-conformation
Binding of core-extended naphthalene diimide (cex-NDI) with hybrid G-quadruplex topology involves no stacking interactions and is featured by the highest fluorescence quantum yield, stacking of cex-NDI on the parallel G-quadruplex topology is associated with intermediate quantum yield, and intercalative stacking of cex-NDI as observed with duplex structures results in the lowest fluorescence quantum yield

Summary

Introduction

G-quadruplex structures are structural motifs of DNA or RNA, whose main unit involves a cyclic planar array of four guanines, which is called G-quartet [1,2,3,4]. A peculiar feature of G-quadruplexes is their remarkable structural polymorphism, which depends on strands stoichiometry and orientation, type of linking loops, and guanine residues conformation. According to the relative orientation of the strands forming the G-quadruplex structure, different topologies, i.e., parallel, antiparallel or hybrid conformations, can be distinguished [1,2]. Studies exploiting antibodies were focused on fixed cells where the structural dynamics of nucleic acids are limited and processes involving G-quadruplexes along with the related downstream cellular pathways cannot be fully understood [20,21,22]

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Results

Conclusion