Simultaneous Binding of Hybrid Molecules Constructed with Dual DNA‐Binding Components to a G‐Quadruplex and Its Proximal Duplex

Abstract

A G-quadruplex (quadruplex) is a nucleic acid secondary structure adopted by guanine-rich sequences and is considered to be relevant to various pharmacological and biological contexts. Although a number of researchers have endeavored to discover and develop quadruplex-interactive molecules, poor ligand designability originating from topological similarity of the skeleton of diverse quadruplexes has remained a bottleneck for gaining specificity for individual quadruplexes. This work reports on hybrid molecules that were constructed with dual DNA-binding components, a cyclic imidazole/lysine polyamide (cIKP), and a hairpin pyrrole/imidazole polyamide (hPIP), with the aim toward specific quadruplex targeting by reading out the local duplex DNA sequence adjacent to designated quadruplexes in the genome. By means of circular dichroism (CD), fluorescence resonance energy transfer (FRET), surface plasmon resonance (SPR), and NMR techniques, we showed the dual and simultaneous recognition of the respective segment via hybrid molecules, and the synergistic and mutual effect of each binding component that was appropriately linked on higher binding affinity and modest sequence specificity. Monitoring quadruplex and duplex imino protons of the quadruplex/duplex motif titrated with hybrid molecules clearly revealed distinct features of the binding of hybrid molecules to the respective segments upon their simultaneous recognition. A series of the systematic and detailed binding assays described here showed that the concept of simultaneous recognition of quadruplex and its proximal duplex by hybrid molecules constructed with the dual DNA-binding components may provide a new strategy for ligand design, enabling targeting of a large variety of designated quadruplexes at specific genome locations.

Ligand Design to Acquire Specificity to Intended G‐Quadruplex Structures

“…Amongt hese, hybrid 3 exhibits a4 .6-fold selectivity against a G4/duplex substrate predicted to be the most likely off target (K D = 3.7 10 À9 versus 17 10 À9 m). [43] The PIP parto ft he hybrid molecules accounts for their selectivity:P IPs are able to recognize aG /C base pair by antiparallel pairing of an imidazole/pyrrole moiety and an A/T or aT /A base pairb ya ntiparallel pairing of P/P pair.Ab-alanine/b-alanine pair reads an A/T or aT /A base pair in the same way as that of aP /P pair. [44] This liganddesign methodology is, in theory,a pplicable to the selective targetingo fabroad varietyo fd esignated quadruplexes in the genome.…”

Duplex‐Guided Refolding into Novel G‐Quadruplex (3+1) Hybrid Conformations

“…[26][27][28] Likewise,t he here reported two novel three-layered (3+ +1) hybrid quadruplexes with one lateral and two propeller loops may be conceivable in G-rich genomic sequences if short single-stranded regions flanking the two outermost functional G-tracts allow for Watson-Crick base pairing.A sd emonstrated by previous studies,t he duplex-quadruplex junction formed may also be au nique interaction site for various ligands. [29,30] On the other hand, the principle of guiding quadruplex folding through additional interactions in duplex extensions may support arational design of an ever increasing toolbox of quadruplex topologies for their use in technological applications.…”

“…[25] In addition to base pairing between overhang sequences, internal hairpin duplexes formed along quadruplex loops have been shown to stabilize quadruplex architectures and seem to be am ore recurrent motif in genomic DNA. [29,30] On the other hand, the principle of guiding quadruplex folding through additional interactions in duplex extensions may support arational design of an ever increasing toolbox of quadruplex topologies for their use in technological applications. [29,30] On the other hand, the principle of guiding quadruplex folding through additional interactions in duplex extensions may support arational design of an ever increasing toolbox of quadruplex topologies for their use in technological applications.…”

Duplex‐gesteuerte Umfaltung in neuartige G‐Quadruplex‐(3+1)‐ Hybridkonformationen

“…Es kann daher vermutet werden, dass temporär gebildete Basenpaarungen zwischen den Überhangbereichen eine Vororientierung der Sequenz bewirken und letztlich mit der Duplexstruktur als kinetischer Falle die Faltung in eine Hybridstruktur begünstigen. [29,30] Aufd er anderen Seite kçnnte das Prinzip einer gesteuerten Quadruplexfaltung durch zusätzliche Duplexerweiterungen ein rationales Design zu einem immer grçßer werdenden Repertoire von Quadruplexstrukturen für deren Verwendung in verschiedenen technischen Anwendungen unterstützen. [25] Außer Basenpaarungen zwischen Überhangsequenzen stabilisieren auch Hairpin-Duplexstrukturen innerhalb der Quadruplex-Loopregionen die G4-Struktur und scheinen zudem häufig in genomischer DNAaufzutreten.…”

“…Wiei nv orherigen Studien gezeigt, kann der Quadruplex-Duplex-Übergang auch als spezifische Bindungsstelle fürv erschiedene Liganden wirken. [29,30] Aufd er anderen Seite kçnnte das Prinzip einer gesteuerten Quadruplexfaltung durch zusätzliche Duplexerweiterungen ein rationales Design zu einem immer grçßer werdenden Repertoire von Quadruplexstrukturen für deren Verwendung in verschiedenen technischen Anwendungen unterstützen.…”