G-quadruplex Research Articles

Dynamics of the 1:2:1 and 1:6:1 C-Myc G-Quadruplexes with the Drude Polarizable Force Field

G-quadruplexes (GQs) are noncanonical nucleic acid structures that form in guanine-rich sequences, importantly gene promoters and telomeres. The guanines within GQs arrange in square, planar tetrads through Hoogsteen hydrogen bonding. These tetrads stack on top of one another to make a highly ordered structure with an electronegative core. Coordinated cations are required to counteract the core's electronegativity and stabilize the GQ. GQs play important roles in gene expression and genomic stability and are functionally relevant in several human proto-oncogenes. One such proto-oncogene, c-myc, is linked to several human cancers such as Burkitt's lymphoma and multiples types of myeloma due to GQ formation within the promoter region of the gene acting as a gene silencer. As such, the c-myc GQ is a novel drug target for cancer therapy. Two solved GQ structures of interest in the c-myc promoter are the 1:2:1 and 1:6:1 GQs, named for differences in central loop length. Loop regions of GQs are known to affect topology, stability, and overall structure. Moreover, studies have shown nucleolin preferentially binds to the 1:6:1 GQ, prompting our present study on the dynamics of c-myc GQs. To analyze the importance of loop sequence on structural dynamics and gain information on this potential drug target, we performed molecular dynamics simulations utilizing the CHARMM36 non-polarizable and the Drude-2017 polarizable force fields (FF). The Drude FF accounts for electronic degrees of freedom via negatively charged particles placed on every heavy atom in the systems and has provided new insights on GQ dynamics and DNA-ion interactions. Our simulations of the 1:2:1 and 1:6:1 GQs demonstrate that polarization is important to model the stability, ion-ion, and ion-DNA interactions that underly GQ dynamics, emphasizing the importance of polarizable molecular dynamics in studying GQs.

Identification and characterization of two conserved G-quadruplex forming motifs in the Nipah virus genome and their interaction with G-quadruplex specific ligands

The G-quadruplex (GQ) motifs are considered as potential drug-target sites for several human pathogenic viruses such as Zika, Hepatitis, Ebola, and Human Herpesviruses. The recent outbreaks of Nipah virus (NiV) in India, the highly fatal emerging zoonotic virus is a potential threat to global health security as no anti-viral drug or vaccine in currently available. Therefore, here in the present study, we sought to assess the ability of the putative G-quadruplex forming sequences in the NiV genome to form G-quadruplex structures and act as targets for anti-viral compounds. Bioinformatics analysis underpinned by various biophysical and biochemical techniques (such as NMR, CD, EMSA, DMS footprinting assay) confirmed the presence of two highly conserved G-quadruplex forming sequences (HGQs) in the G and L genes of NiV. These genes encode the cell attachment glycoprotein and RNA-dependent RNA polymerase, respectively and are essential for the virus entry and replication within the host cell. It remains possible that stabilization of these HGQs by the known G-quadruplex binding ligands like TMPyP4 and Braco-19 represents a promising strategy to inhibit the expression of the HGQ harboring genes and thereby stop the viral entry and replication inside the host cell. Accordingly, we report for the first time, that HGQs in Nipah virus genome are targets for G-quadruplex specific ligands; therefore, could serve as potential targets for anti-viral therapy.

Intrinsic “Turn-On” Aptasensor Detection of Ochratoxin A Using Energy-Transfer Fluorescence

Ochratoxin A (OTA) is an intrinsically fluorescent phenolic mycotoxin that contaminates a wide range of food products and is a serious health threat to animals and humans. An OTA binding aptamer (OTABA) that folds into an antiparallel G-quadruplex (GQ) in the absence and presence of target OTA has been incorporated into a vast variety of aptasensor platforms for OTA detection. The development of a simple, aptamer-based approach would allow for detection of the toxin without the use of complex analytical instrumentation, which has been the gold standard for OTA detection thus far. However, to date none of the aptasensor platforms have utilized the natural fluorescence of the phenolic toxin for detection. Herein, we report that OTA binding to OTABA involves pi-stacking interactions that leads to GQ-to-toxin energy transfer (ET), which affords a 'turn-on' fluorescence self-signaling platform in which the emission of the aptamer‒target complex is enhanced in comparison to the free toxin alone. Selective excitation of the GQ‒OTA complex at 256 nm leads to a 4-fold enhancement in OTA fluorescence. The GQ‒OTA ET detection platform boasts a limit of detection ~ 2 ng/mL, which is comparable to a previously demonstrated FRET immunoassay platform for OTA detection, and displays excellent OTA selectivity and recovery from red wine samples.

Prion protein transcription is auto-regulated through dynamic interactions with G-quadruplex motifs in its own promoter

Cellular prion protein (PrP) misfolds into an aberrant and infectious scrapie form (PrPSc) that lead to fatal transmissible spongiform encephalopathies (TSEs). Association of prions with G-quadruplex (GQ) forming nucleic acid motifs has been reported, but implications of these interactions remain elusive. Herein, we show that the promoter region of the human prion gene (PRNP) contains two putative GQ motifs (Q1 and Q2) that assume stable, hybrid, intra-molecular quadruplex structures and bind with high affinity to PrP. Here, we investigate the ability of PrP to bind to the quadruplexes in its own promoter. We used a battery of techniques including SPR, NMR, CD, MD simulations and cell culture-based reporter assays. Our results show that PrP auto-regulates its expression by binding and resolving the GQs present in its own promoter. Furthermore, we map this resolvase-like activity to the N-terminal region (residues 23–89) of PrP. Our findings highlight a positive transcriptional-translational feedback regulation of the PRNP gene by PrP through dynamic unwinding of GQs in its promoter. Taken together, our results shed light on a yet unknown mechanism of regulation of the PRNP gene. This work provides the necessary framework for a plethora of studies on understanding the regulation of PrP levels and its implications in prion pathogenesis.

Recognition of G-quadruplex topology through hybrid binding with implications in cancer theranostics.

The selective recognition and imaging of oncogene specific G-quadruplex (GQ) structures holds great promise in the development of diagnostic therapy (theranostics) for cancer and has been challenging due to their structural dynamics and diversity. We report selective recognition of GQ by a small molecule through unique hybrid loop stacking and groove binding mode with turn on far-red fluorescence response and anticancer activity demonstrating the potential implications for GQ-targeted cancer theranostics.Methods: Biophysical investigation reveal the turn on far-red emission property of TGP18 for selective recognition of GQ. In cellulo studies including DNA damage and oxidative stress evaluation guided us to perform in vitro (3D spheroid) and in vivo (xenograft mice model) anti-cancer activity, and tumor tissue imaging to assess the theranostic potential of TGP18.Results: Neocuproine-based far-red turn on fluorescence probe TGP18 shows GQ-to-duplex selectivity and specifically recognizes BCL-2 GQ with high affinity through a unique hybrid binding mode involving loop-stacking and groove interactions. Our study reveals that the selective recognition originating from the distinct loop structure of GQ that alters the overall probe interaction and binding affinity. TGP18 binding to anti-apoptotic BCL-2 GQ ablates the pro-survival function and elicit anti-cancer activity by inducing apoptosis in cancer cells. We deciphered that inhibition of BCL-2 transcription synergized with signaling cascade of nucleolar stress, DNA damage and oxidative stress in triggering apoptosis signaling pathway.Conclusion: Intervention of GQ mediated lethality by TGP18 has translated into anti-cancer activity in both in vitro 3D spheroid culture and in vivo xenograft models of lung and breast cancer with superior efficacy for the former. In vivo therapeutic efficacy supplemented with tumor 3D spheroid and tissue imaging potential define the role of TGP18 in GQ-targeted cancer theranostics.

Ratiometric fluorescent sensing of the parallel G-quadruplex produced by PS2.M: implications for K+ detection.

Fluorescent ligands that selectively bind to a specific G-quadruplex (GQ) topology (antiparallel, hybrid or parallel) are highly sought after for aptasensor development and nanodevice construction. The coumarin-benzothiazole hybrid (BnBtC) is an internal charge transfer (ICT) ratiometric fluorescent probe, which displays two well-resolved emission bands at ∼450 nm for the coumarin component and ∼650 nm for the ICT band. The red ICT emission of BnBtC displays turn-on responses to protic solvent polarity and upon binding GQ structures, especially those produced by the hemin binding aptamer (PS2.M). In the present work, BnBtC was found to exhibit enhanced ICT emission upon binding the parallel GQ topology of PS2.M that is selectively produced in the presence of K+. This ability to discriminate K+ from other cationic metal ions through a turn-on ratiometric fluorescent response demonstrates the potential utility of the BnBtC probe for biosensor applications.

De novo post-SELEX optimization of a G-quadruplex DNA aptamer binding to marine toxin gonyautoxin 1/4

Ligand-binding aptamers obtained by SELEX (Systematic Evolution of Ligands by EXponential enrichment) often have low affinity or/and specificity, and post-SELEX optimization is usually needed. Due to experimental difficulty in determining three-dimensional (3D) structures of aptamer-ligand complexes, there are few structure-guided methods for rational post-SELEX optimization. Here, we employed a de novo optimization approach to engineer high-affinity variants for a G-quadruplex (GQ) aptamer (GO18-T-d) that specifically binds to marine toxin gonyautoxin 1/4 (GTX1/4). First, temperature-dependent modeling was carried out to predict the atomic structure of GO18-T-d. Then, to identify key bases for the optimization, spontaneous binding simulations were performed to reveal the complex structure of GO18-T-d with GTX1/4. Finally, binding energy analysis was conducted to evaluate the designed variants for high affinity. We predicted that GO18-T-d has the typical parallel GQ topology, consistent with circular dichroism (CD) measurements. Our simulations showed that the 5′-end of GO18-T-d hinders the GTX1/4 movement toward the binding pocket, leading to a designed variant that removes the first 5 nucleotides at the 5′-end. Microscale thermophoresis (MST) experiments verified that the binding affinity of the engineered aptamer increases by ~20 folds. Thus, this study not only provides a high-affinity variant of GO18-T-d, but also suggests that our computational approach is useful for the structure-guided optimization of GQ aptamers.

Two novel fan-shaped trinuclear Pt(ii) complexes act as G-quadruplex binders and telomerase inhibitors.

Two new trinuclear Pt(ii) complexes {[Pt(dien)]3(tib)}(NO3)6 (1) and {[Pt(dpa)]3(tib)}(NO3)6 (2) (dien: diethylenetriamine, dpa: bis-(2-pyridylmethyl)amine, tib: 1,3,5-tris(1H-imidazol-1-yl)benzene) have been designed, synthesized, characterized and applied to a series of biochemical studies. We found that both of the Pt(ii) complexes exhibited much better selectivity for human telomeric G-quadruplex sequence than promoter G-quadruplexes (c-kit, c-myc, and bcl2) or duplex DNA. Both complexes displayed comparative stability and affinity towards human telomeric G-quadruplex by the studies from surface plasmon resonance, fluorescence resonance energy transfer and polymerase chain reaction stop assays. The circular dichroism indicated that both complexes could induce and stabilize anti-parallel G-quadruplex structures. Molecule docking presented that Pt(ii) complex intercalated into the large groove of human telomeric G-quadruplex (PDB ID: 143D). Furthermore, telomeric repeat amplification protocol assays quantitatively evaluated the inhibition of telomerase activity caused by the Pt(ii) complexes. The obtained IC50 values of 6.41 ± 0.042 μM and 2.67 ± 0.035 μM for 1 and 2, respectively, exhibited strong telomerase inhibitions. All results suggest that such fan-shaped trinuclear Pt(ii) complexes are effective and selective G-quadruplex binders, as well as strong telomerase inhibitors. This study provides insight into the development of human telomeric G-quadruplex targeted anticancer drugs based on the metal complex.

Substituted adenine quartets: interplay between substituent effect, hydrogen bonding, and aromaticity.

Adenine, one of the components of DNA/RNA helices, has the ability to form self-organizing structures with cyclic hydrogen bonds (A4), similar to guanine quartets. Here, we report a computational investigation of the effect of substituents (X = NO2, Cl, F, H, Me, and NH2) on the electronic structure of 9H-adenine and its quartets (A4-N1, A4-N3, and A4-N7). DFT calculations were used to show the relationships between the electronic nature of the substituents, strength of H-bonds in the quartets, and aromaticity of five- and six-membered rings of adenine. We demonstrated how the remote substituent X modifies the proton-donating properties of the NH2 group involved in the H-bonds within quartets and how the position of the substituent and its electronic nature affect the stability of the quartets. We also showed the possible changes in electronic properties of the substituent and aromaticity of adenine rings caused by tetramer formation. The results indicate that the observed relationships depend on the A4 type. Moreover, the same substituent can both strengthen and weaken intermolecular interactions, depending on the substitution position.

Properties of the excited electronic states of guanine quartet complexes with alkali metal cations

G-quartets are supra-molecular structures that consist of four guanine molecules connected by eight hydrogen bonds. They are additionally stabilized by metal cations. In this contribution, the excited states of G-quartet and its complexes with lithium, sodium and potassium were studied by employing time-dependent density functional theory. The findings indicate that vertical excitations from the optimized ground state involve transitions from several bases, whereas excitations from the optimized lowest excited state include transitions from one base. The charge-transfer character of these states was analyzed. It was shown that the cations are able to modify positions of the maxima of the fluorescence spectra of the complexes.

G-quadruplexes in fibrotic scars may open new therapeutic avenues for wound healing.

Read the Original article at https://doi.org/10.1002/1873-3468.13658.

Same fold, different properties: polarizable molecular dynamics simulations of telomeric and TERRA G-quadruplexes.

DNA and RNA sequences rich in guanine can fold into noncanonical structures called G-quadruplexes (GQs), which exhibit a common stem structure of Hoogsteen hydrogen-bonded guanine tetrads and diverse loop structures. GQ sequence motifs are overrepresented in promoters, origins of replication, telomeres, and untranslated regions in mRNA, suggesting roles in modulating gene expression and preserving genomic integrity. Given these roles and unique aspects of different structures, GQs are attractive targets for drug design, but greater insight into GQ folding pathways and the interactions stabilizing them is required. Here, we performed molecular dynamics simulations to study two bimolecular GQs, a telomeric DNA GQ and the analogous telomeric repeat-containing RNA (TERRA) GQ. We applied the Drude polarizable force field, which we show outperforms the additive CHARMM36 force field in both ion retention and maintenance of the GQ folds. The polarizable simulations reveal that the GQs bind bulk K+ ions differently, and that the TERRA GQ accumulates more K+ ions, suggesting different ion interactions stabilize these structures. Nucleobase dipole moments vary as a function of position and also contribute to ion binding. Finally, we show that the TERRA GQ is more sensitive than the telomeric DNA GQ to water-mediated modulation of ion-induced dipole-dipole interactions.

Cationic porphyrins as destabilizer of a G-quadruplex located at the promoter of human MYH7 β gene

G-quadruplex (GQ) architecture is adopted by guanine rich sequences, present throughout the eukaryotic genome including promoter locations and telomeric ends. The in vivo presence indicates their involvement and role in various biological processes. Various small ligands have been developed to interact and stabilize/destabilize G-quadruplex structures. Cationic porphyrins are among the most studied ligands, reported to bind and stabilize G-quadruplexes. Herein, we report the recognition and destabilization of a parallel G-quadruplex by porphyrins (TMPyP3 and TMPyP4). This G-quadruplex forming 23-nt G-rich sequence is in the promoter region of Human Myosin Heavy Chain β gene (MYH7β). Presence of various putative regulatory sequence elements (TATA Box, CCAAT, SP-1) located in the vicinity of this quadruplex motif, highlight its regulatory implications. Biophysical methods as Circular Dichroism Spectroscopy, UV-Absorption Spectroscopy, UV-Thermal Denaturation and Fluorescence Spectroscopy (steady as well as Time Resolved) have been used for studying the interaction and binding parameters. It is proposed that porphyrins have a destabilizing effect on the G-quadruplexes with parallel topology and a stronger binding specifically via intercalation mode is needed to cause destabilization. The study deals with better understanding and insights of DNA-Drug interactions in biological systems.Communicated by Ramaswamy H. Sarma

N-methyl mesoporphyrin IX as a highly selective light-up probe for G-quadruplex DNA.

N-methyl mesoporphyrin IX (NMM) is a water-soluble, non-symmetric porphyrin with excellent optical properties and unparalleled selectivity for G-quadruplex (GQ) DNA. G-quadruplexes are non-canonical DNA structures formed by guanine-rich sequences. They are implicated in genomic stability, longevity, and cancer. The ability of NMM to selectively recognize GQ structures makes it a valuable scaffold for designing novel GQ binders. In this review, we survey the literature describing the GQ-binding properties of NMM as well as its wide utility in chemistry and biology. We start with the discovery of the GQ-binding properties of NMM and the development of NMM-binding aptamers. We then discuss the optical properties of NMM, focusing on the light-switch effect — high fluorescence of NMM induced upon its binding to GQ DNA. Additionally, we examine the affinity and selectivity of NMM for GQs, as well as its ability to stabilize GQ structures and favor parallel GQ conformations. Furthermore, a portion of the review is dedicated to the applications of NMM-GQ complexes as biosensors for heavy metals, small molecules (e.g. ATP and pesticides), DNA, and proteins. Finally and importantly, we discuss the utility of NMM as a probe to investigate the roles of GQs in biological processes.

Real-time Monitoring Excitation Dynamics of Human Telomeric Guanine Quadruplexes: Effect of Folding Topology, Metal Cation, and Confinement by Nanocavity Water Pool.

Guanine(G)-rich human telomeric (HT) DNA repeats, crucial to maintenance of genome stability, readily form G-quadruplexes (GQs) with various folding topologies. Research on excitation dynamics of HT-GQs is, however, scarce. Herein, we report a femtosecond time-resolved fluorescence coupled with transient absorption investigation on HT-GQ with the basket-type structure in Na+ solution. The result unveils an unusual multichannel nonradiative mechanism dominated by states with varying character of charge transfer lasting ten and hundreds of picoseconds, accounting altogether for an overwhelming ∼85% of the overall deactivation involving proton transfer. Our comparative study shows that encapsulating the GQ in nanocavity water pool or changing it into hydrid-type topologies with the presence of K+ ions alter differently energies and lifetimes of these states, yet without affecting the nature of the electronic excitation involved. The finding of this work underscores a leading role of structural rigidity in regulating the interplay with the microenvironment of photoexcited monomolecularly folded HT-GQs.

G-quadruplex structures trigger RNA phase separation.

Liquid–liquid phase separation plays an important role in a variety of cellular processes, including the formation of membrane-less organelles, the cytoskeleton, signalling complexes, and many other biological supramolecular assemblies. Studies on the molecular basis of phase separation in cells have focused on protein-driven phase separation. In contrast, there is limited understanding on how RNA specifically contributes to phase separation. Here, we described a phase-separation-like phenomenon that SHORT ROOT (SHR) RNA undergoes in cells. We found that an RNA G-quadruplex (GQ) forms in SHR mRNA and is capable of triggering RNA phase separation under physiological conditions, suggesting that GQs might be responsible for the formation of the SHR phase-separation-like phenomenon in vivo. We also found the extent of GQ-triggered-phase-separation increases on exposure to conditions which promote GQ. Furthermore, GQs with more G-quartets and longer loops are more likely to form phase separation. Our studies provide the first evidence that RNA can adopt structural motifs to trigger and/or maintain the specificity of RNA-driven phase separation.

Targeting eIF4A Using MZ-735 Potently Induces Cell Death in Lymphoma Cells and Rapidly Represses mRNA Translation at the Global Level and in C-MYC and Other Oncogenes

Introduction: Dysregulated C-MYC signaling is associated with the pathogenesis and poor survival in aggressive lymphomas such as diffuse large B cell lymphoma (DLBCL). Despite intense investigation, direct inhibitors of C-MYC have not yet advanced to the clinic. Various structural elements, such as TOP (terminal oligopyrimidine tract) motif, guanine quartet (CGG) 4 motif, and polypurine sequences (AGAGAG), have been shown to impart high level of dependence on eukaryotic initiation factor 4F (eIF4F). The eIF4F is comprised of 3 subunits, including the mRNA 5ʹ-cap-binding subunit eIF4E, the large scaffolding subunit eIF4G, and the RNA helicase subunit eIF4A. In the current project, we have investigated targeting eIF4A as a novel therapeutic strategy in lymphoma. Pateamine A (PatA) is a natural product originally isolated from a New Zealand marine sponge, Mycale sp. DMDA-PatA, a first generation PatA analogue, has demonstrated in vivo activity in xenograft cancer models. DMDA-PatA acts by decreasing the interaction between eIF4A and eIF4G that specifically inhibits cap dependent translation. MZ-735 is a recently developed PatA analogue that is more potent than DMDA-PatA and less protein bound. Materials and Methods: MZ-735 was synthesized at the CPRIT Synthesis and Drug-Lead Discovery Laboratory at Baylor University. Cytotoxicity was evaluated in diffuse large B cell and Mantle cell lymphoma cell lines using CellTiter-Glo (Promega®). Western blot analysis was performed for the expressions of oncogenes. Global translation was determined using surface sensing of translation (SUnSET) assay and Polysome profile followed by Western blotting and qPCR. RNA sequencing (RNA-seq) was carried out on drug versus dimethyl sulfoxide (DMSO)-treated cells in DLBCL and MCL. An unbiased proteomics analysis was performed using TMT mass spectrometry. Omics data were analyzed by gene set enrichment analysis (GSEA) and Virtual Inference of Protein-activity by Enriched Regulon (VIPER) analysis. Results: MZ-735 were studied in 13 lymphoma cell lines. MZ-735 exhibited concentration- and time-dependent cytotoxicity in lymphoma cell lines at low nanomolar concentrations and showed minimal toxicity in normal PBMC cells. MZ-735 potently inhibits polysome formation, global protein synthesis and inhibits oncogene such as C-Myc and Cyclin D1 in DLBCL and MCL cells. An unbiased quantitative proteomics analysis was carried out to identify the immediate consequence of MZ-735 during a short exposure time of 3 hours. More than 5000 proteins were detected, of which 132 proteins were upregualated and 119 proteins were down regulated by more than twofold after MZ-735 treatment (P < 0.05; fold change, >2). Enrichment analysis suggested that Cancer hallmark proteins such as E2F target proteins, G2/M check point proteins and mitotic spindle proteins were highly enriched among these depleted proteins and the most affected downregulated pathways are cell cycle, cell division and chromosome segregation pathway. We are currently conducting RNAseq to determine whether the mRNA level change in the same or opposite direction as the protein level for any given gene across the genome. Furthermore, we will conduct proteomics and RNAseq in samples treated for longer time points, as the short exposure (3h) favors discovery of depleted proteins with very short half-lives. Conclusion: These results demonstrate that the PatA analogue MZ-735 can potently inhibit the translational apparatus in lymphoma. MZ-735 inhibits metabolic pathways required for cancer progression, leading to growth arrest and apoptosis in the lymphoma cells. MZ-735 and other eIF4A inhibitors represent a promising new class of anti-neoplastic agents, which may be particularly useful for silencing undruggable oncogenes such as C-MYC. Deep insights into the translational responses to eIF4A inhibitors at the single gene level will inform future development of this class of anti-neoplastic agents. Disclosures O'Connor: Allos Therapeutics: Consultancy; Acetylon Pharma: Other: Travel expenses, Research Funding; Celgene: Research Funding; Millenium: Consultancy, Honoraria, Other: Travel expenses, Research Funding; Mundipharma: Consultancy, Honoraria, Other: Travel expenses, Research Funding; Novartis: Consultancy, Honoraria; Roche: Research Funding; Seattle Genetics, Inc.: Consultancy, Other: Travel expenses, Research Funding; Spectrum Pharma: Consultancy, Other: Travel expenses, Research Funding.

Development of a highly sensitive label-free DNA based fluorescent sensor for cisplatin detection

Cisplatin, or cis-diamminedichloroplatinum (II), is a platinum-based chemotherapeutic drug. It is widely used against many types of cancer. However, there are various side effects in patients taking cisplatin. The side effects are also found in chemotherapists or health workers exposed to contaminating cisplatin in workplace. Therefore, there is a need for sensors for cisplatin contamination detection. This work reports the development of a highly sensitive fluorescent sensor using single stranded ssDNA as a sensing element. It contains consecutive guanine bases to allow an efficient cisplatin binding. Then, an enzyme-free DNA amplification technique called catalyzed hairpin assembly (CHA) was employed to increase the sensor sensitivity. It comprises two hairpin DNAs (H1 and H2) which are in meta-stable states and can form a hybrid only in the presence of the cisplatin sensing ssDNA described above. H1 is designed such that after the hybrid formation, a newly exposed overhang forms a G-quadruplex (GQ). The GQ formation can be monitored by mixing a GQ binding dye called Thioflavin T (ThT) in the solution. ThT fluorescence quantum yield is negligible if ThT is free in solution but increases over 1000 folds when bound to GQ. The formation of H1:H2 hybrid and ThT-GQ complex is disrupted if cisplatin binds to the sensing ssDNA. Therefore, the decrease in ThT fluorescence signifies the increase in cisplatin concentration and vice versa. The developed sensor capable to detect the cisplatin in the range of 500 – 7500 nM and the limit of detection is 182 nM. The merit of this novel sensor is that it is label-free therefore economical yet highly sensitive. It is expected that the sensor can help monitor cisplatin contamination in chemotherapeutic workplaces.

Streamlining effects of extra telomeric repeat on telomeric DNA folding revealed by fluorescence-force spectroscopy.

A human telomere ends in a single-stranded 3′ tail, composed of repeats of T2AG3. G-quadruplexes (GQs) formed from four consecutive repeats have been shown to possess high-structural and mechanical diversity. In principle, a GQ can form from any four repeats that are not necessarily consecutive. To understand the dynamics of GQs with positional multiplicity, we studied five and six repeats human telomeric sequence using a combination of single molecule FRET and optical tweezers. Our results suggest preferential formation of GQs at the 3′ end both in K+ and Na+ solutions, with minor populations of 5′-GQ or long-loop GQs. A vectorial folding assay which mimics the directional nature of telomere extension showed that the 3′ preference holds even when folding is allowed to begin from the 5′ side. In 100 mM K+, the unassociated T2AG3 segment has a streamlining effect in that one or two mechanically distinct species was observed at a single position instead of six or more observed without an unassociated repeat. We did not observe such streamlining effect in 100 mM Na+. Location of GQ and reduction in conformational diversity in the presence of extra repeats have implications in telomerase inhibition, T-loop formation and telomere end protection.

Applications of guanine quartets in nanotechnology and chemical biology

Guanine and related nucleobases such as guanosine, deoxyguanosine and isoguanosine are notable molecular tools for designing functional supramolecular assemblies. This popularity originates in their ability to self-assemble via a unique topological pluralism — as isolated nucleobases, discrete macrocyclic quartets and virtually infinite linear ribbons — that endows them with a considerable functional versatility. Many programmes have been launched to fine-tune the chemical properties of guanine derivatives, to make them usable under different experimental conditions, such as in organic or aqueous environments, and responsive to external stimuli, such as ionic strength, pH, light or temperature. These strategies aim to translate the chemical information encoded in a basic guanine unit into programmable, higher-order supramolecular architectures. Spectacular results have been recently obtained in various chemical fields, from supramolecular chemistry to chemical biology, from soft matter to catalysis. In this Review, we detail these advances and demonstrate how these multidisciplinary investigations cast a bright light on the diversity that guanines, synthetic guanines and related nucleobases uniquely offer in terms of both structure and function. The propensity of guanine and its derivatives to assemble into guanine quartets makes them a privileged tool in the design of functional supramolecular assemblies. This Review describes the production of such functional suprastructures and their applications in nanotechnology, soft matter and chemical biology.