DNA In Aqueous Solution Research Articles

Generation of Guanine–Thymidine Cross-Links in DNA by Peroxynitrite/Carbon Dioxide

Nitrosoperoxycarbonate derived from the combination of carbon dioxide and peroxynitrite is an important chemical mediator of inflammation. In aqueous solutions, it rapidly decomposes to the reactive species CO(3)(•-) and (•)NO(2) radicals that are known to initiate the selective oxidation and nitration of guanine in DNA. We have previously demonstrated that the reactions of carbonate radical anions with guanine in 2'-deoxyoligoribonucleotides generate a previously unknown intrastrand cross-linked guanine-thymine product G*-T* with a covalent bond between the C8 (G*) and the thymine N3 (T*) atoms (Crean Nucleic Acids Res. 2008, 36, 742-755). In this work, we demonstrate that G*-T* cross-linked products are also formed when peroxynitrite (0.1 mM) reacts with native DNA in aqueous solutions (pH 7.5-7.7) containing 25 mM carbon dioxide/bicarbonate, in addition to the well-known nitration/oxidation products of guanine such as 8-nitroguanine (8-nitro-G), 5-guanidino-4-nitroimidazole (NIm), 8-oxo-7,8-dehydroguanine (8-oxo-G), and spiroiminodihydantoin (Sp). The yields of these products, after enzymatic digestion with P1 nuclease and alkaline phosphatase to the nucleotide level and reversed phase HPLC separation, were compared with those obtained with the uniformly, isotopically labeled (15)N,(13)C-labeled 2'-deoxy oligoribonucleotides 5'-dGpT and 5'-dGpCpT. The d(G*pT*) and d(G*-T*) cross-linked products derived from the di- and trioligonucleotides, respectively, were used as standards for identifying the analogous lesions in calf thymus DNA by isotope dilution LC-MS/MS methods in the selected reaction monitoring mode. The NIm and 8-nitro-G are the major products formed (∼0.05% each), and lesser amounts of 8-oxo-G (∼0.02%) and d(G*pT*) and d(G*-T*) enzymatic digestion products (∼0.002% each) were found. It is shown that the formation of d(G*pT*) enzyme digestion product can arise only from intrastrand cross-links, whereas d(G*-T*) can arise from both interstrand and intrastrand cross-linked products.

DNA hydrogels: New functional soft materials

AbstractDeoxyribonucleic acid (DNA) hydrogel is a network of crosslinked DNA strands swollen in aqueous solutions. The crosslinks may be physical or chemical, such as the hydrogen bonds or ethylene glycol units, respectively, connecting the strands belonging to different double‐helical DNA molecules. As DNA network strands in the hydrogels exhibit properties similar to those of the individual DNA molecules, such soft materials are a good candidate to make use of the characteristics of DNA such as coil‐globule transition, biocompatibility, selective binding, and molecular recognition. Physical DNA hydrogels with an elastic modulus in the order of megapascals can be prepared by subjecting semidilute aqueous solutions of DNA to successive heating–cooling cycles between below and above the melting temperature of DNA. Chemical DNA hydrogels can be prepared by connecting the amino groups on the nucleotide bases through covalent bonds to form a three‐dimensional DNA network in aqueous solutions. In this article, we summarize the preparation strategies of DNA hydrogels with a wide range of tunable properties. © 2011 Wiley Periodicals, Inc. J Polym Sci Part B: Polym Phys, 2011

Preparation of a [Ru(bpy)2(dppz)]2+-Intercalated DNA Cast Film Using a Self-Standing Method and Its Luminescence Tuning by Cu2+ Ions and EDTA

In this correspondence, we report on the first preparation of [Ru(bpy)(2)(dppz)](2+)-intercalated (bpy = 2,2'-bipyridine; dppz = dipyrido[3,2-a:2',3'-c]phenazine) DNA films on an indium-tin oxide surface via a solution-based self-standing strategy, carried out by the direct mixing of aqueous solutions of both anionic DNA and cationic metallointercalator at a molar ratio of 5:6. The luminescence of a [Ru(bpy)(2)(dppz)](2+)-intercalated DNA cast film is studied and found to show excellent tunable characteristics by Cu(2+) ions and ethylenediaminetetraacetic acid addition.

Cholesteric and Nematic Liquid Crystalline Phase Behavior of Double-Stranded DNA Stabilized Single-Walled Carbon Nanotube Dispersions

The first lyotropic cholesteric single-walled carbon nanotube (SWNT) liquid crystal phase was obtained by dispersing SWNTs in an aqueous solution of double-stranded DNA (dsDNA). Depending on the dispersion methodology, the polydomain nematic phase previously reported for other lyotropic carbon nanotube dispersions could also be obtained. The phase behavior and dispersion microstructure were affected by the relative concentrations of dsDNA and SWNT and whether small bundles were removed prior to concentrating the dispersions. This readily controlled phase behavior opens new routes for producing SWNT films with controlled morphology.

Viscosity of aqueous DNA solutions determined using dynamic light scattering

Viscosity is a key parameter for characterising the behaviour of liquids and their flow. It is, however, difficult to measure precisely, reproducibly and accurately for aqueous solutions on a micro-litre volume scale, which is what is usually needed for biological samples. We report the development of a new method for measuring dynamic viscosity by measuring dynamic light scattering (DLS) data for a range of particles of well-defined size. Most applications of DLS involve determining particle size for samples of known viscosity. We inverted the usual protocol and endeavoured to determine viscosity for samples of known particle size. Viscosity measurements for water and aqueous solutions of calf thymus DNA made using DLS were compared with those from a U-tube viscometer. The styrene particles, frequently used as particle size standards, gave unsatisfactory results for our DNA samples as did C-6 derivatized silica and positively charged amino polystyrene microspheres. However, negatively charged carboxylate polystyrene microspheres particles readily gave accurate viscosity measurements over a range of temperatures (0-100 °C). The sample volume required depends on the cuvette used to measure DLS, but can be performed with samples sizes ranging from 40 to 3000 μL. The sample can then be recovered for subsequent experiments. The DLS method is simple to perform at different temperatures and provides data of accuracy significantly above that of a U-tube viscometer. Our results also indicate a way forward to account accurately for solution viscosity in the normal applications of DLS to particle size determination by including the appropriate non-interacting particles as an internal standard.

Heuristic Correlation of Rheological Measurements and Electrochemical Impedance Spectroscopy (EIS) of the DNA/Water System

In this work, it is reported viscosity, density, sound velocity and electrochemical impedance spectroscopy (EIS) measurements of calf thymus DNA in aqueous solutions, in order to investigate the overlapping C* and entanglement concentrations Ce. These critical concentrations demark the conditions at which start appearing interactions and entanglements between DNA molecules in solution. EIS has been proposed as new alternative analytical techniques to obtain C* and Ce in polyelectrolyte solutions. A new parameter the characteristic time (τc) was defined, and it is obtained from a new type of log-log Bode plot using the crossover frequency (τc=1/ωc) of the real (Z') and imaginary (Z") components when they are plotted as a function of the frequency. Preliminary results show similar values C* and Ce by EIS to those obtained from rheological, densimetry and sound velocity measurements.

Electric dichroism transients of aqueous solutions of DNA

In this work we develop a theory of reduced electric linear dichroism transients of DNAfragments in aqueous solution. The DNA fragments are modelled as rigid ‘bent-rod molecules’(BRM) with the following physical parameters: electric charge, electric polarizabilitytensors and hydrodynamical ones, and the average transition probability tensor permolecule. In order to study the growth and decay of electric dichroism transients, theorientational distribution function of the molecules is needed. This function is obtained bysolving the time-dependent Fokker–Planck equation in the presence of a low electric fieldE, using a perturbation method and the Fourier method with time-dependent coefficients. Inour calculations the origin of the coordinate system is the mass centre of the BRM. Withrespect to this centre, the electric dipole moment of the molecule is zero. The developedtheory adequately explains the experimental results. We show that the theoretical approachused in this work is equivalent to the one applied in the Brownian dynamics simulationwork performed by Porschke and co-workers. We also analyse the effect of a possibleelectric dipole moment on the transients of the reduced electric linear dichroism in DNAbent fragments.

Study of DNA damage with a new system for irradiation of samples in a nuclear reactor

In this paper, we report results of a quantitative analysis of the effects of neutrons on DNA, and, specifically, the production of simple and double breaks of plasmid DNA in aqueous solutions with different concentrations of free-radical scavengers. The radiation damage to DNA was evaluated by electrophoresis through agarose gels. The neutron and gamma doses were measured separately with thermoluminescent detectors. In this work, we have also demonstrated usefulness of a new system for positioning and removing samples in channel BH#3 of the IEA-R1 reactor at the Instituto de Pesquisas Energéticas e Nucleares (Brazil) without necessity of interrupting the reactor operation.

Effect of divalent cations on DNA condensed on the surface of rigid cationic membrane

It has been indicated in previous literature reports that divalent cations cannot directly condense DNA in aqueous solution. However, several studies have shown that divalent metal ions can induce a further condensation of DNA originally adsorbed on the surfaces of fluid cationic membranes. In this article, we examine the effect of divalent metal ions (Ca 2+ and Ni 2+) on DNA condensed on the surfaces of a rigid cationic membrane formed by the mixture of cholesteryl 3-N-(dimethyl–aminoethyl) carbamate (DC-Chol) and 1,2-di(cis-9-octadecenoyl)-sn-glycero-3-phosphocholine (DOPC). Our results demonstrate that the divalent cations exert an electrostatic screening effect between DNA, where the in-plane spacing between DNA ( d DNA) confined between the lipid bilayers gradually decreases with increasing cation concentration when the concentration exceeds a threshold value. The collapse transition, where d DNA drops discontinuously at a critical cation concentration, observed previously for the fluid membrane is not identified here, indicating the critical role of membrane flexibility in the two-dimensional condensation of DNA.

The Effect of Electroporation Type Pulsed Electric Fields on DNA in Aqueous Solution

Electroporation is a physical phenomenon in which pulsed electric fields applied across a cell produce transient (reversible) or permanent (irreversible) permeabilization of the cell membrane. Irreversible electroporation is an important method of sterilization in the food industry and it is becoming an important minimally invasive tissue ablation technique in medicine. Motivated by recent observations of apoptosis like marker stains in irreversibly electroporated cells we performed a study on the effects of electroporation type electric pulses on the integrity of naked DNA in solution. Using gel electrophoresis analyses we show that pulses of the irreversible electroporation type have the ability to affect the naked DNA in solution. It is found that some electric parameters that lead to cell death by irreversible electroporation also cause changes in the naked DNA exposed to the same procedure. Our analysis tentatively suggests that some electroporation type electric pulses cause nicks in the DNA molecule. Therefore, it is possible that the mechanisms of cell death in irreversible electroporation also include damages to the DNA. However, this work did not investigate the possible effects of electroporation induced electrode corrosion byproducts, such as Al(3+) ions on DNA integrity; which should be also studied in the future. In general, since electroporation phenomena based applications are widely used in medicine and biotechnology, the current study suggests that further research into the effects of electroporation type electric pulses on the DNA are warranted.

Structure reactivity relationship in the reaction of DNA guanyl radicals with hydroxybenzoates

In DNA, guanine bases are the sites from which electrons are most easily removed. As a result of hole migration to this stable location on guanine, guanyl radicals are major intermediates in DNA damage produced by the direct effect of ionizing radiation (ionization of the DNA itself and not through the intermediacy of water radicals). We have modeled this process by employing gamma irradiation in the presence of thiocyanate ions, a method which also produces single electron oxidized guanyl radicals in plasmid DNA in aqueous solution. The stable products formed in DNA from these radicals are detected as strand breaks after incubation with the FPG protein. When a phenolic compound is present in the solution during gamma irradiation, the formation of guanyl radical species is decreased by electron donation from the phenol to the guanyl radical. We have quantified the rate of this reaction for four different phenolic compounds bearing carboxylate substituents as proton acceptors. A comparison of the rates of these reactions with the redox strengths of the phenolic compounds reveals that salicylate reacts ca. 10-fold faster than its structural analogs. This observation is consistent with a reaction mechanism involving a proton coupled electron transfer, because intra-molecular transfer of a proton from the phenolic hydroxyl group to the carboxylate group is possible only in salicylate, and is favored by the strong 6-membered ring intra-molecular hydrogen bond in this compound.

SPECTROSCOPIC AND PHYSICOCHEMICAL METHODS FOR STUDYING THE INTERACTION OF METALLOPORPHYRIN WITH DNA

In recent years studies on the interaction of porphyrin with DNA have received much attention because of the importance in DNA-probing and photodynamic therapy of cancer. A variety of spectroscopic methods, e.g. NMR, ESR, Mössbauer, UV-visible absorption, circular dichroism (CD), magnetic circular dichroism (MCD), IR and Raman spectroscopy, have been employed for studying interactions between porphyrin and DNA. Of these spectroscopic methods, only a few instrumental analytical techniques applicable to an aqueous buffer solution of DNA have been particularly developed to investigate porphyrin-DNA interactions. On the other hand, a number of physicochemical methods, e.g. gel electrophoresis, melting temperature measurements and hydrodynamic methods such as viscosity and sedimentation measurements, have been also used for determining the binding modes of porphyrin to DNA. The present lecture will focus on the application of visible absorption, CD and MCD spectroscopy as well as melting temperature and viscosity measurements to studies of porphyrin-DNA interactions.

Low-frequency spectroscopy of four-photon scattering of laser radiation in aqueous solutions of biopolymers

Four-photon scattering (FPS) spectroscopy is used to record rotational resonances of H2O and H2O2 molecules in aqueous solutions of DNA and denatured DNA within a range of ±10 cm-1 with a spectral resolution of 3 GHz. The resonance contribution of rotational transitions of these molecules in solutions was found to be considerably larger than that in distilled water. This fact is interpreted as a manifestation of the specific properties of a hydrate layer at the interface between water and DNA or denatured DNA molecules. An analysis of the FPS spectra shows that the concentration of H2O2 molecules in the hydrate layer of the DNA molecule increases threefold after denaturation. In addition, the FPS spectra of aqueous solutions of α-chymotrypsin protein with concentrations of 0-20 mg cm-3 were measured in the spectral range of ±7 cm-1. It is found that the velocity of hypersound in the protein aqueous solution, which is measured by the shift of the Mandelstam—Brillouin scattering spectrum components, is a cubic function of the concentration and reaches 3000 m/s at 20 mg/cm3.

Detecting DNA Folding with Nanocapillaries

We demonstrate for the first time the detection of the folding state of double-stranded DNA in nanocapillaries with the resistive pulse technique. We show that glass capillaries can be pulled into nanocapillaries with diameters down to 45 nm. We study translocation of lambda -DNA which is driven by an electrophoretic force through the nanocapillary. The resulting change in ionic current indicates the folding state of single lambda -DNA molecules. Our experiments prove that nanocapillaries are suitable for label-free analysis of DNA in aqueous solutions and viable alternatives to solid-state nanopores made by silicon nanotechnology.

β-Carboline alkaloids bind DNA

β-Carboline alkaloids present in Peganum harmala ( harmal) have recently drawn attention due to their antitumor activities. The mechanistic studies indicate that beta-carboline derivatives inhibit DNA topoisomerases and interfere with DNA synthesis. They interact with DNA via both groove binding and intercalative modes and cause major DNA structural changes. The aim of this study was to examine the interactions of five β-carboline alkaloids (harmine, harmane, harmaline, harmalol and tryptoline) with calf-thymus DNA in aqueous solution at physiological conditions, using constant DNA concentration (6.25 mM) and various alkaloids/polynucleotide (phosphate) ratios of 1/240, 1/160, 1/80, 1/40, 1/20, 1/10, 1/5, 1/2 and 1/1. Fourier transform infrared (FTIR) and UV–visible spectroscopic methods were used to determine the ligand binding modes, the binding constants, and the stability of alkaloids–DNA complexes in aqueous solution. Spectroscopic evidence showed major binding of alkaloids to DNA with overall binding constants of K harmine − DNA = 3.44 × 10 7 M −1, K harmane − DNA = 1.63 × 10 5 M −1, K harmaline − DNA = 3.82 × 10 5 M −1, K harmalol − DNA = 6.43 × 10 5 M −1 and K tryptoline − DNA = 1.11 × 10 5 M −1. The affinity of alkaloids–DNA binding is in the order of harmine > harmalol > harmaline > harmane > tryptoline. No biopolymer secondary structural changes were observed upon alkaloid interaction and DNA remains in the B-family structure in these complexes.

Toward a full structural characterization of G-quadruplex DNA in aqueous solution: Molecular dynamics simulations of four G-quadruplex molecules

A systematic study on guanine (G) quadruplex DNA flexibility was performed by using extended explicit molecular dynamics (MD) simulations on four quadruplex molecules containing one dimeric and three monomeric G-quadruplexes with the most used and recent modified versions of AMBER force fields (parm99 and parmbsc0). The detailed analyses of general structure and basic structural parameters were done for the G-DNA structures. The results indicate that parmbsc0 provides a slightly better description for the G-DNA stems than parm99, and the situation is the opposite for the loops. Many backbone torsions located in the non-canonical regions are not retained in both simulations. Most of the glycosidic torsion angles of loop bases deviate largely from the experimental values. Many phase angles of pseudorotation of the sugar rings are transformed to other puckers. The transformation of structural parameters given by each force field is useful as direction for further improving force field of polynucleotide structures.

Synthesis of a new β-naphthothiazole monomethine cyanine dye for the detection of DNA in aqueous solution

Novel monomethine cyanine dye (MC) derived from beta-naphthothiazole and benzothiazole has been prepared and characterized by (1)H and (13)C NMR, FTIR, ESIMS, elemental analyses, absorption and fluorescence spectroscopy. The dye was conveniently synthesized by the condensation of two sulfate heterocyclic quaternary salts. The interaction between calf thymus DNA (ct-DNA) in tris(hydroxymethyl)aminomethane-HCl (Tris-HCl) aqueous buffer solution and MC has been studied with spectral fluorescence method. The binding constant value has been determined by fluorescence titration of MC with ct-DNA concentrations. The result obtained is consistent with an intercalative binding interaction between MC and ct-DNA. Compared with ethidium bromide (EB), MC showed a huge fluorescence enhancement upon mixing with ct-DNA.

Properties of DNA + Dipropylsulfoxide or Dibutylsulfoxide + Water Ternary Solutions

Aqueous solutions of DNA in the presence of dipropylsulfoxide (DPSO) and of dibutylsulfoxide (DBSO) have been studied using UV-vis spectroscopy and density measurements. It has been shown that significant thermal denaturation of DNA occurs when DPSO or DBSO are present. The results obtained were compared with those obtained previously with dimethylsulfoxide (DMSO) and diethylsulfoxide (DESO). It is suggested that the lowering of the transition enthalpy with increasing length of the carbon chain of the dialkylsulfoxides is due to significance contribution from hydrophobic interactions.

DNA Compaction on Histone Mimics Prepared from Silica Nanoparticles

Abstract Silica nanoparticles, the surface of which was modified by a 3-aminopropylsilyl group, compacted gene-size DNA in aqueous solutions in the presence of Na+ or K+. The DNA-compaction efficiency of histone-size (10 nm diameter) nanoparticles is greater than that of large (100 nm) nanoparticles under similar experimental conditions.

Length Distribution of Single‐Walled Carbon Nanotubes in Aqueous Suspension Measured by Electrospray Differential Mobility Analysis

The first characterization of the length distribution of single-walled carbon nanotubes (SWCNT) dispersed in a liquid by electrospray differential mobility analysis (ES-DMA) is presented. Although an understanding of geometric properties of SWCNTs, including length, diameter, aspect ratio, and chirality, is essential for commercial applications, rapid characterization of nanotube length distributions remains challenging. Here the use of ES-DMA to obtain length distributions of DNA-wrapped SWCNTs dispersed in aqueous solutions is demonstrated. Lengths measured by ES-DMA compare favorably with those obtained from multiangle light scattering, dynamic light scattering, field flow fractionation with UV/vis detection, and atomic force microscopy, validating ES-DMA as a technique to measure SWCNTs of <250 nm in length. The nanotubes are previously purified and dispersed by wrapping with oligomeric DNA in aqueous solution and centrifuging to remove bundles and amorphous carbon. These dispersions are particularly attractive due to their amenability to bulk processing, ease of storage, high concentration, compatibility with biological and high-throughput manufacturing environments, and for their potential applications ranging from electronics and hydrogen-storage vessels to anticancer agents.