Dinuclear Ruthenium(II) Complexes That Induce and Stabilise G‐Quadruplex DNA

A series of dinuclear ruthenium(II) complexes were synthesised, and the complexes were determined to be new highly selective compounds for binding to telomeric G‐quadruplex DNA. The interactions of these complexes with telomeric G‐quadruplex DNA were studied by using circular dichroism (CD) spectroscopy, fluorescence resonance energy transfer (FRET) melting assays, isothermal titration calorimetry (ITC) and molecular modelling. The results showed that the complexes 1 , 2 and 4 induced and stabilised the formation of antiparallel G‐quadruplexes of telomeric DNA in the absence of salt or in the presence of 100 mM K⁺‐containing buffer. Furthermore, complexes 1 and 2 strongly bind to and effectively stabilise the telomeric G‐quadruplex structure and have significant selectivity for G‐quadruplex over duplex DNA. In comparison, complex 3 had a much lesser effect on the G‐quadruplex, suggesting that possession of a suitably sized plane for good π–π stacking with the G‐quadruplets is essential for the interaction of the dinuclear ruthenium(II) complexes with the G‐quadruplex. Moreover, telomerase inhibition by the four complexes and their cellular effects were studied, and complex 1 was determined to be the most promising inhibitor of both telomerase and HeLa cell proliferation.     

A Twisting Electronic Nanoswitch Made of DNA

Single‐stranded DNAs and RNAs that are rich in the nucleobase guanine form four‐stranded G‐quadruplexes, which are held together by hydrogen‐bonded guanine quartets. In aqueous solution, both DNA duplexes and G‐quadruplexes are modest conductors of electrical charge. A tight, topologically constrained DNA construct called twDNA is now reported, in which a core of four guanine‐rich single strands structurally and electronically links together four DNA double helices. The addition and removal of K⁺ or Sr²⁺ cations promote alternative conformers of twDNA, which have strikingly distinct electronic properties. Unlike DNA mechano‐electronic switches that require large conformational changes, twDNA requires only modest twisting/untwisting structural attenuations to achieve electronic switching.     

Ion‐Selective Formation of a Guanine Quadruplex on DNA Origami Structures

DNA origami nanostructures are a versatile tool that can be used to arrange functionalities with high local control to study molecular processes at a single‐molecule level. Here, we demonstrate that DNA origami substrates can be used to suppress the formation of specific guanine (G) quadruplex structures from telomeric DNA. The folding of telomeres into G‐quadruplex structures in the presence of monovalent cations (e.g. Na⁺ and K⁺) is currently used for the detection of K⁺ ions, however, with insufficient selectivity towards Na⁺. By means of FRET between two suitable dyes attached to the 3′‐ and 5′‐ends of telomeric DNA we demonstrate that the formation of G‐quadruplexes on DNA origami templates in the presence of sodium ions is suppressed due to steric hindrance. Hence, telomeric DNA attached to DNA origami structures represents a highly sensitive and selective detection tool for potassium ions even in the presence of high concentrations of sodium ions.     

Involvement of Long‐Lived Intermediate States in the Complex Folding Pathway of the Human Telomeric G‐Quadruplex

The energy landscapes of human telomeric G‐quadruplexes are complex, and their folding pathways have remained largely unexplored. By using real‐time NMR spectroscopy, we investigated the K⁺‐induced folding of the human telomeric DNA sequence 5′‐TTGGG(TTAGGG)₃A‐3′. Three long‐lived states were detected during folding: a major conformation (hybrid‐1), a previously structurally uncharacterized minor conformation (hybrid‐2), and a partially unfolded state. The minor hybrid‐2 conformation is formed faster than the more stable hybrid‐1 conformation. Equilibration of the two states is slow and proceeds via a partially unfolded intermediate state, which can be described as an ensemble of hairpin‐like structures.     

Formation of Human Telomeric G-quadruplex Structures Induced by the Quaternary Benzophenanthridine Alkaloids:Sanguinarine,Nitidine,and Chelerythrine

The ligands which can facilitate the formation and stabilize G‐quadruplex structures have attracted enormous attention due to their potential ability of inhibiting the telomerase activity and halting tumor cell proliferation. It is noteworthy that the abilities of the quaternary benzophenanthridine alkaloids (QBAs), the very important G‐quadruplex binders, in inducing the formation of human telomeric DNA G‐quadruplex structures, have not been reported. Herein, the interaction between single‐strand human telomeric DNA and three QBAs: Sanguinarine (San), Nitidine (Nit) and Chelerythrine (Che), has been investigated. Although these molecules are very similar in structure, they exhibit significantly different abilities in inducing oligonucleotide d(TTAGGG)₄ (HT4) to specific G‐quadruplex structures. Our experimental results indicated that the best ligand San could convert HT4 into antiparallel G‐quadruplex structure completely, followed by Nit, which could transform to mixed‐type or hybrid G‐quadruplex structure partially, whereas Che could only transform to antiparallel G‐quadruplex structure in small quantities. The relative QBAs' inducing abilities as indicated by the CD data are in the order of San>Nit>Che. Further investigation revealed that the G‐quadruplex structures from HT4 induced by QBAs are of intramolecular motif. And only sequences with certain length could be induced by QBAs because of their positive charges which could not attract short chain DNA molecules to close to each other and form intermolecular G‐quadruplex. In addition, the factors that affect the interaction between HT4 and QBAs were discussed. It is proposed that the thickness of the molecular frame and the steric hindrance are the primary reasons why the subtle differences in QBAs' structure lead to their remarkable differences in inducing the formation of the G‐quadruplex structures.     

Synthesis of bis-indole carboxamides as G-quadruplex stabilizing and inducing ligands

The design and synthesis of a series of bis‐indole carboxamides with varying amine containing side chains as G‐quadruplex DNA stabilising small molecules are reported. Their interactions with quadruplexes have been evaluated by means of Förster resonance energy transfer (FRET) melting analysis, UV/Vis spectroscopy, circular dichroism spectroscopy and molecular modelling studies. FRET analysis indicates that these ligands exhibit significant selectivity for quadruplex over duplex DNA, and the position of the carboxamide side chains is of paramount importance in G‐quadruplex stabilisation. UV/Vis titration studies reveal that bis‐indole ligands bind tightly to quadruplexes and show a three‐ to fivefold preference for c‐kit2 over h‐telo quadruplex DNA. CD studies revealed that bis‐indole carboxamide with a central pyridine ring induces the formation of a single, antiparallel, conformation of the h‐telo quadruplex in the presence and absence of added salt. The chirality of h‐telo quadruplex was transferred to the achiral ligand (induced CD) and the formation of a preferred atropisomer was observed.     

Specific recognition of human telomeric G-quadruplex DNA with small molecules and the conformational analysis by ESI mass spectrometry and circular dichroism spectropolarimetry

Electrospray ionization mass spectrometry (ESI-MS) was utilized to investigate the binding affinity and stoichiometry of small molecules with human telomeric G-quadruplex DNA. The binding-affinity order obtained for the (AGGGTT)(4) quadruplex was: Tel01>ImImImbetaDp>PyPyPygammaImImImbetaDp. The specific binding of Tel01 and PyPyPygammaImImImbetaDp in one system consisting of human telomeric G-quadruplex and duplex DNA was observed directly for the first time. This revealed that PyPyPygammaImImImbetaDp has a binding specificity for the duplex DNA, whereas Tel01 selectively recognizes the G-quadruplex DNA. Moreover, both ESI-MS and circular dichroism (CD) spectra indicated that Tel01 favored the formation and stabilization of the antiparallel G-quadruplex, and a structural transition of the (AGGGTT)(4) sequence from a coexistence of parallel and antiparallel G-quadruplexes to a parallel G-quadruplex induced by annealing.     

Environment‐Sensitive Probes Containing a 2,6‐Diethynylpyridine Motif for Fluorescence Turn‐On Detection and Induction of Nanoarchitectures of Human Telomeric Quadruplex

Bis(phenylethynyl)pyridylcarboxamides with amide side chains at the para position of the NH₂ group possess strong solvatochromic properties compared with the meta analogues. Fluorescence binding titrations show that these probes exhibit remarkable fluorescence turn‐on responses upon interacting with the human telomeric G‐quadruplex (h‐TELO). Förster resonance energy transfer melting analysis shows the high selectivity of these probes for h‐TELO over duplex DNA. Isothermal titration calorimetry, as well as UV/Vis and fluorescence spectroscopy studies, show that the meta analogue has a twofold binding affinity for h‐TELO over the para analogue. The noncovalent interaction of these small‐molecule probes with h‐TELO has been used to regulate the assembly of novel supramolecular nanoarchitectures.     

Formation and stability of G-quadruplexes self-assembled from guanine-rich strands

Electrospray ionization mass spectrometry (ESI-MS) was utilized to investigate the formation and stability of G-quadruplexes. For the 15 6-nt oligonucleotides tested, ESI-MS indicated that formation of a parallel tetramer quadruplex requires at least four continuous guanines in the 6-nt sequence. In addition, the G-rich strands prefer to employ "self-association" in the formation of the G-quadruplex rather than hybridized integration, and the thermodynamic-stability order of these three G-quadruplexes is Q(2)>Q(1)>Q(3).     

Programmed Self‐Assembly of a Quadruplex DNA Nanowire

The ability to produce, reproducibly and systematically, well‐defined quadruplex DNA nanowires through controlled rational design is poorly understood despite potential utility in structural nanotechnology. The programmed hierarchical self‐assembly of a long four‐stranded DNA nanowire through cohesive self‐assembly of GpC and CpG “sticky” ends is reported. The encoding of bases within the quadruplex stem allows for an uninterrupted π‐stacking system with rectilinear propagation for hundreds of nanometers in length. The wire is mechanically stable and features superior nuclease resistance to double‐stranded DNA. The study indicates the feasibility for programmed assembly of uninterrupted quadruplex DNA nanowires. This is fundamental to the systematic investigation of well‐defined DNA nanostructures for uses in optoelectronic and electronic devices as well as other structural nanotechnology applications.     

Multitasking Water‐Soluble Synthetic G‐Quartets: From Preferential RNA–Quadruplex Interaction to Biocatalytic Activity

Natural G‐quartets, a cyclic and coplanar array of four guanine residues held together through a Watson–Crick/Hoogsteen hydrogen‐bond network, have received recently much attention due to their involvement in G‐quadruplex DNA, an alternative higher‐order DNA structure strongly suspected to play important roles in key cellular events. Besides this, synthetic G‐quartets (SQ), which artificially mimic native G‐quartets, have also been widely studied for their involvement in nanotechnological applications (i.e., nanowires, artificial ion channels, etc.). In contrast, intramolecular synthetic G‐quartets (iSQ), also named template‐assembled synthetic G‐quartets (TASQ), have been more sparingly investigated, despite a technological potential just as interesting. Herein, we report on a particular iSQ named ^PNADOTASQ, which demonstrates very interesting properties in terms of DNA and RNA interaction (notably its selective recognition of quadruplexes according to a bioinspired process) and catalytic activities, through its ability to perform peroxidase‐like hemin‐mediated oxidations either in an autonomous fashion (i.e., as pre‐catalyst for TASQzyme reactions) or in conjunction with quadruplex DNA (i.e., as enhancing agents for DNAzyme processes). These results provide a solid scientific basis for TASQ to be used as multitasking tools for bionanotechnological applications.