Guanine tetraplex topology of human telomere DNA is governed by the number of (TTAGGG) repeats

Secondary structures of the G-rich strand of human telomere DNA fragments G(3)(TTAG(3))(n), n = 1–16, have been studied by means of circular dichroism spectroscopy and PAGE, in solutions of physiological potassium cation concentrations. It has been found that folding of these fragments into tetraplexes as well as tetraplex thermostabilities and enthalpy values depend on the number of TTAG(3) repeats. The suggested topologies include, e.g. antiparallel and parallel bimolecular tetraplexes, an intramolecular antiparallel tetraplex, a tetraplex consisting of three parallel chains and one antiparallel chain, a poorly stable parallel intramolecular tetraplex, and both parallel and antiparallel tetramolecular tetraplexes. G(3)(TTAG(3))(3) folds into a single, stable and very compact intramolecular antiparallel tetraplex. With an increasing repeat number, the fragment tetraplexes surprisingly are ever less thermostable and their migration and enthalpy decrease indicate increasing irregularities or domain splitting in their arrangements. Reduced stability and different topology of lengthy telomeric tails could contribute to the stepwise telomere shortening process

The guanine-rich fragile X chromosome repeats are reluctant to form tetraplexes

Using circular dichroism spectroscopy, UV absorption spectroscopy and polyacrylamide gel electrophoresis, we studied conformational properties of guanine-rich DNA strands of the fragile X chromosome repeats d(GGC)(n), d(GCG)(n) and d(CGG)(n), with n = 2, 4, 8 and 16. These strands are generally considered in the literature to form guanine tetraplexes responsible for the repeat expansion. However, we show in this paper that the repeats are reluctant to form tetraplexes. At physiological concentrations of either Na(+) or K(+) ions, the hexamers and dodecamers associate to form homoduplexes and the longer repeats generate homoduplexes and hairpins. The tetraplexes are rarely observed being relatively most stable with d(GGC)(n) and least stable with d(GCG)(n). The tetraplexes are exclusively formed in the presence of K(+) ions, at salt concentrations higher than physiological, more easily at higher than physiological temperatures, and they arise with extremely long kinetics (even days). Moreover, the capability to form tetraplexes sharply diminishes with the oligonucleotide length. These facts make the concept of the tetraplex appearance in this motif in vivo very improbable. Rather, a hairpin of the fragile X repeats, whose stability increases with the repeat length, is the probable structure responsible for the repeat expansion in genomes

Influence of additional bases and of a redox marker on the structure of the Thrombin Binding Aptamer

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On the interaction between [Ru(NH3)6]3+ and the G-quadruplex forming thrombin binding aptamer sequence

The interaction between the thrombin binding aptamer (TBA), a G-quadruplex forming DNA sequence, andthe electroactive hexaammineruthenium(III) cation has been studied by electrochemical methods and circulardichroism spectroscopy. When TBA is immobilised on a gold surface in a typical aptasensor configuration,the [Ru(NH3)6]3+ cation can be bound to the electrode surface through its interaction with the TBA sequence.This interaction is strong enough to enable the rutheniumcomplex to remain at the surface when the electrode isimmersed in an electrolyte free of [Ru(NH3)6]3+, meaning that the complex does not diffuse back into thesolution. A stoichiometry of 2 [Ru(NH3)6]3+ per TBA strand has been determined, indicating that the interactiondiffers from the conventional, non-specific electrostatic charge compensation, forwhich a 5 to 1 ratiowould be expected between the triply charged cation and the 15 bases sequence. It is shown that this interactiontakes place not only at the surface, but also when both TBA and hexaammineruthenium(III) aredissolved in solution. Under such conditions, a similar stoichiometry of 2 [Ru(NH3)6]3+ per TBA strandhas been evidenced by two independent methods, namely circular dichroism spectroscopy and differentialpulse voltammetry.SCOPUS: ar.jinfo:eu-repo/semantics/publishe

Specific interaction between a redox marker and G quartets

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Electrochemical and circular dichroism spectroscopic evidence of two types of interaction between [Ru(NH3)6]3+ and an elongated thrombin binding aptamer G-quadruplex

In the field of electrochemical biosensing based on DNA probes immobilised at electrode surfaces the [Ru(NH3)6]3+ redox marker is commonly used for quantitative characterisation and/or detection purposes. Although it is generally assumed that the interactions between the hexaammineruthenium(III) cation and DNA are solely of electrostatic nature, and thus non-specific in essence, it is shown herein that this is not the case for a sequence containing a G-quadruplex part. The present work investigates the interactions of [Ru(NH3)6]3+ with an oligonucleotide (21-mer) consisting of the thrombin binding aptamer sequence (a G-quadruplex) elongated by a 5’-overhang of six nucleotides. Combining circular dichroism and electrochemical analyses by differential pulse voltammetry, alternating current voltammetry and cyclic voltammetry, it is demonstrated that the overall interaction stoichiometry (number of [Ru(NH3)6]3+ bound to one DNA strand) is lower than the 7 [Ru(NH3)6]3+ per strand expected for a total charge compensation. Our results indicate that two types of interaction take place between [Ru(NH3)6]3+ and the elongated TBA sequence: one is the conventional electrostatic binding of the positively charged redox marker with the negatively charged phosphate groups, while the other is connected with the G-quadruplex part of the sequence. This latter interaction influences the DNA conformation in solution and involves 2 [Ru(NH3)6]3+ per strand, both in solution and with DNA immobilised at gold electrodes. In this configuration, the electron transfer rate constant is evaluated at 37 ± 5 s-1 on the basis of a Laviron plot.info:eu-repo/semantics/publishe

Elongated thrombin binding aptamer: a G-quadruplex cation-sensitive conformational switch.

Aptamer-based biosensors offer promising perspectives for high performance, specific detection of proteins. The thrombin binding aptamer (TBA) is a G-quadruplex-forming DNA sequence, which is frequently elongated at one end to increase its analytical performances in a biosensor configuration. Herein, we investigate how the elongation of TBA at its 5' end affects its structure and stability. Circular dichroism spectroscopy shows that TBA folds in an antiparallel G-quadruplex conformation with all studied cations (Ba(2+), Ca(2+), K(+), Mg(2+), Na(+), NH(4)(+), Sr(2+) and the [Ru(NH(3))(6)](2+/3+) redox marker) whereas other structures are adopted by the elongated aptamers in the presence of some of these cations. The stability of each structure is evaluated on the basis of UV spectroscopy melting curves. Thermal difference spectra confirm the quadruplex character of all conformations. The elongated sequences can adopt a parallel or an antiparallel structure, depending on the nature of the cation; this can potentially confer an ion-sensitive switch behavior. This switch property is demonstrated with the frequently employed redox complex [Ru(NH(3))(6)](3+), which induces the parallel conformation at very low concentrations (10 equiv per strand). The addition of large amounts of K(+) reverts the conformation to the antiparallel form, and opens interesting perspectives for electrochemical biosensing or redox-active responsive devices.Journal ArticleResearch Support, Non-U.S. Gov'tFLWINSCOPUS: ar.jinfo:eu-repo/semantics/publishe

CD spectra of GTTAG (left) and TAGTTAGT (right) measured at 25°C (dots) in 1 mM Na phosphate plus 0

<p><b>Copyright information:</b></p><p>Taken from "Guanine tetraplex topology of human telomere DNA is governed by the number of (TTAGGG) repeats"</p><p>Nucleic Acids Research 2005;33(18):5851-5860.</p><p>Published online 12 Oct 2005</p><p>PMCID:PMC1253834.</p><p>© The Author 2005. Published by Oxford University Press. All rights reserved</p>3 mM EDTA, pH 7 immediately after thermal denaturation. Measurements in 10 mM potassium phosphate plus 0.15 M KCl were carried out (left): immediately (dashes) and 1, 3 and 10 days (from the thinnest to the thickest line) after K addition. The samples were kept and measured at 25°C; (right): after two days keeping at 0°C and measured at 0°C (dashes) and 25°C (dash–dots) and after three days keeping at 25°C and measured at 25°C (thick line). The thin full line spectra in both panels correspond to the samples kept for two days in 10 mM Na phosphate plus 0.15 M NaCl at 0°C and measured at 0°C. Inset: UV absorption spectra of GTTAG in 10 mM K phosphate plus 0.15 M KCl at 91°C (dots) and 25°C (solid line). The sketch shows the antiparallel and parallel bimolecular tetraplexes of TAGTTAGT. The balls in the sketch refer to the oligonucleotide 5′ ends

IFI16 Preferentially Binds to DNA with Quadruplex Structure and Enhances DNA Quadruplex Formation.

Interferon-inducible protein 16 (IFI16) is a member of the HIN-200 protein family, containing two HIN domains and one PYRIN domain. IFI16 acts as a sensor of viral and bacterial DNA and is important for innate immune responses. IFI16 binds DNA and binding has been described to be DNA length-dependent, but a preference for supercoiled DNA has also been demonstrated. Here we report a specific preference of IFI16 for binding to quadruplex DNA compared to other DNA structures. IFI16 binds to quadruplex DNA with significantly higher affinity than to the same sequence in double stranded DNA. By circular dichroism (CD) spectroscopy we also demonstrated the ability of IFI16 to stabilize quadruplex structures with quadruplex-forming oligonucleotides derived from human telomere (HTEL) sequences and the MYC promotor. A novel H/D exchange mass spectrometry approach was developed to assess protein interactions with quadruplex DNA. Quadruplex DNA changed the IFI16 deuteration profile in parts of the PYRIN domain (aa 0-80) and in structurally identical parts of both HIN domains (aa 271-302 and aa 586-617) compared to single stranded or double stranded DNAs, supporting the preferential affinity of IFI16 for structured DNA. Our results reveal the importance of quadruplex DNA structure in IFI16 binding and improve our understanding of how IFI16 senses DNA. IFI16 selectivity for quadruplex structure provides a mechanistic framework for IFI16 in immunity and cellular processes including DNA damage responses and cell proliferation

Long-read sequencing technology indicates genome-wide effects of non-B DNA on polymerization speed and error rate.

DNA conformation may deviate from the classical B-form in ∼13% of the human genome. Non-B DNA regulates many cellular processes; however, its effects on DNA polymerization speed and accuracy have not been investigated genome-wide. Such an inquiry is critical for understanding neurological diseases and cancer genome instability. Here, we present the first simultaneous examination of DNA polymerization kinetics and errors in the human genome sequenced with Single-Molecule Real-Time (SMRT) technology. We show that polymerization speed differs between non-B and B-DNA: It decelerates at G-quadruplexes and fluctuates periodically at disease-causing tandem repeats. Analyzing polymerization kinetics profiles, we predict and validate experimentally non-B DNA formation for a novel motif. We demonstrate that several non-B motifs affect sequencing errors (e.g., G-quadruplexes increase error rates), and that sequencing errors are positively associated with polymerase slowdown. Finally, we show that highly divergent G4 motifs have pronounced polymerization slowdown and high sequencing error rates, suggesting similar mechanisms for sequencing errors and germline mutations