B-form Of DNA Research Articles

Assessment of A- to B- DNA Transitions Utilizing the Drude Polarizable Force Field.

In addition to the well-characterized B-form of DNA, duplex DNA can adopt various conformations, such as A or Z-DNA. Though less common, these structures can be induced biologically through protein or ligand interactions or experimentally with niche environmental conditions, such as high salt concentrations or in mixed water-ethanol. Reproducing these alternate structures through molecular dynamics simulations in recent years has been quite challenging with the currently available force fields, simulation techniques, and time scales. In this study, the Drude polarizable force field is tested for its ability to facilitate transitions between A-DNA and B-DNA or maintain A-DNA. Though transitions away from B-DNA were observed in high concentrations of ethanol, the resulting structures had hybrid properties taken from both B-DNA and A-DNA structures. This was also true for A-DNA in ethanol, which lost some of the A-DNA properties that it was expected to maintain. When B-DNA was tested in high salt environments, the resulting B-DNA structures showed no distinguishable differences with the increasing salt concentrations tested. These results with the Drude FF and recent results with additive force fields suggest that at present the current additive and polarizable force fields do not facilitate a complete transition between B- to A-DNA conformations under the conditions simulated. At present, the Drude FF favors A-B DNA hybrid structures when simulated in nonphysiological conditions.

Synthesis, characterization and multi-spectroscopic DNA/HSA interaction studies of synthetic human Follicle-Stimulating Hormone Beta 33–53 peptide conjugated PEGylated graphene oxide nanoparticles

The objective of the current study was to synthesize, characterize and explore the interaction of PEGylated graphene oxide (pGO) and synthetic human Follicular stimulating hormone β 33–53 peptide conjugated PEGylated graphene oxide nanoparticles (pGO-FSH) with human serum albumin (HSA) and calf thymus DNA (CT-DNA). The pGO/ pGO-FSH nanoparticles were synthesized using a modified Hummer’s method, and the FSH peptide was conjugated through a maleimide crosslinking reaction. Synthesized nanoparticles were then characterized using techniques like FT-IR, UV–Visible absorbance, CD and Raman spectroscopy, and XRD and TGA. Morphological and particle size analysis was studied using SEM, TEM, DLS, and zeta potential measurements. The presence of FSH β 33–53 peptide was confirmed qualitatively and quantitatively using CD spectroscopy and Bradford’s assay. Binding studies of pGO/pGO-FSH nanoparticles with HSA and DNA were carried out using biophysical techniques. The complex formation between pGO/pGO-FSH nanoparticles and HSA was revealed by UV absorbance spectroscopy, and the observed fluorescence quenching was confirmed by steady-state fluorescence spectroscopy. Time-resolved fluorescence quenching studies have shown that dynamic quenching plays an important role in binding HSA with pGO/pGO-FSH nanoparticles. However, structurally no significant changes were observed in the native structure of HSA upon binding with pGO/pGO-FSH nanoparticles suggesting that the latter did not induce any structural distortions together, confirmed by DSC, FT-IR, and CD spectroscopy experimental findings. Binding constants and thermodynamic parameters calculated using double logarithmic and Van’t Hoff plots suggested weak and moderate binding affinity along with the involvement of hydrophobic and hydrogen bonding interactions between HSA and pGO/pGO-FSH nanoparticles, respectively. UV absorbance and fluorescence spectroscopy have revealed that pGO/pGO-FSH nanoparticles interact with DNA by binding at the minor groove region. These findings were further confirmed by DNA melting and viscosity studies. CD and FT-IR spectroscopy studies have shown no changes in the helical structure of B-form of DNA, thereby emphasizing the groove-binding nature of pGO/pGO-FSH nanoparticles. The obtained results are useful in further considering the potentiality of pGO-FSH nanoparticles as drug-delivery systems for in vivo applications, especially to target ovarian cancer.

Development of Salmon Sperm DNA/Regenerated Silk Bio-Based Films for Biomedical Studies on Human Keratinocyte HaCaT Cells under Solar Spectrum.

In this study, we fabricated adhesive patches from silkworm-regenerated silk and DNA to safeguard human skin from the sun's rays. The patches are realized by exploiting the dissolution of silk fibers (e.g., silk fibroin (SF)) and salmon sperm DNA in formic acid and CaCl2 solutions. Infrared spectroscopy is used to investigate the conformational transition of SF when combined with DNA; the results indicated that the addition of DNA provides an increase in the SF crystallinity. UV-Visible absorption and circular dichroism spectroscopy showed strong absorption in the UV region and the presence of B-form of DNA once dispersed in the SF matrix, respectively. Water absorption measurements as well as thermal dependence of water sorption and thermal analysis, suggested the stability of the fabricated patches. Biological results on cellular viability (MTT assay) of keratinocyte HaCaT cells after exposures to the solar spectrum showed that both SF and SF/DNA patches are photo-protective by increasing the cellular viability of keratinocytes after UV component exposure. Overall, these SF/DNA patches promise applications in wound dressing for practical biomedical purposes.

The structure of DNA fragments: Quantum-chemical modelling

In this review, we analyze and systematize our computational studies of the nucleic acid duplex formations and thermodynamic stability under the different factors of investigation. The proposed structural models of mini-helix contains N nucleobase pairs (N = 3-5); QM structural data suggest that the helical conformations of mini-helix adopt geometrical parameters comparable to those of natural A- and B-DNA forms under specific conditions as micro hydration and charge compensation. The gas-phase models adopt non regular conformations between the helical form and a ladder form.. The natural helical shape of DNA mini-helix is stabilized by the presence of counterions or by explicit micro-hydration of the major and minor groves. The presence of aqueous solution is shown as a minor factor for the helical shape formation. The studies are performed at the level of density functional theory.

Detection of Z-DNA Structures in Supercoiled Genome.

Z-DNAs are nucleic acid secondary structures that form within a special pattern of nucleotides and are promoted by DNA supercoiling. Through Z-DNA formation, DNA encodes information by dynamic changes in its secondary structure. A growing body of evidence indicates that Z-DNA formation can play a role in gene regulation; it can affect chromatin architecture and demonstrates its association with genomic instability, genetic diseases, and genome evolution. Many functional roles of Z-DNA are yet to be discovered highlighting the need for techniques to detect genome-wide folding of DNA into this structure. Here, we describe an approach to convert linear genome into supercoiled genome sponsoring Z-DNA formation. Applying permanganate-based methodology and high-throughput sequencing to supercoiled genome allows genome-wide detection of single-stranded DNA. Single-stranded DNA is characteristic of the junctions between the classical B-form of DNA and Z-DNA. Consequently, analysis of single-stranded DNA map provides snapshots of the Z-DNA conformation over the whole genome.

A novel approach for identification of mirror repeats within the Engrailed Homeobox-1 gene of Xenopus tropicalis

Background: Repetitive sequences constitute the major portion of genomic DNA in most of the organisms and are responsible for variation in DNA structure, function, etc., These sequences also have the potential to adopt various noncanonical DNA structures. Methods: By using a swift, manual approach mirror repeats has been identified within the complete engrailed homeobox-1 gene (en-1) of X. tropicalis. Another tool Non- B DNA motif search was also deployed for comparative analysis. Results: A total of 166 mirror repeats were identified within the complete en-1 gene of X. tropicalis. The similar sequences were also searched among the genome of different organisms such as Xenopus laevis, Caenorhabditis elegans, Drosophila melanogaster, etc., Conclusion: To the best of our knowledge, it was novel identification of mirror repeats in the engrailed-1 gene of X. tropicalis. Few of these sequences may adopt various noncanonical B-DNA forms and are potent sites for mutation and recombination events.

Interaction of calf thymus DNA and glucose-based gemini cationic surfactants with different spacer length: A spectroscopy and DLS study

The interactions between calf thymus DNA and a series of glucose-based cationic gemini surfactants 1a-1c with different spacer length, n = 4, 6 and 8, were studied by UV absorption, fluorescence spectroscopy, circular dichroism, FT-IR, dynamic light scattering and zeta potential measurements. The results showed that all the surfactants could interact with DNA efficiently. On addition of increasing concentration of the surfactants, UV absorption hypochromicity with insignificant blue shift were observed, until the DNA signal disappeared. The surfactant 1c was more efficient in the reduction of absorption intensity of DNA. According to the fluorescence quenching experiments by ethidium bromide exclusion, 1c exhibited the highest binding properties, with the binding constant at 3.25 × 108 L·mol−1. The spectroscopy study indicated that the surfactants bound with the DNA by a non-intercalative mode, mainly electrostatic interaction between the positively charged headgroups of the surfactants and negatively charged phosphate groups of DNA at low concentration, and the hydrophobic interaction among the alkyl chains at high concentration. The conformation of DNA during the interaction process could be kept B-form of DNA. For 1c, the DNA molecules can be compacted to about 103 nm in hydrodynamic diameter at 0.2 mM, while the minimum sizes of DNA were 140 nm and 133 nm, respectively, in the presence of 1a and 1b. The impact of the cationic gemini surfactants on the DNA compaction and condensation would shed light on their potential applications in gene delivery.

A spectroscopic deciphering of the differential interaction behavior of alkaloid drugs with native B-DNA and protonated DNA

The present study demonstrates a detailed spectroscopic investigation on the differential extent and modes of interaction of two alkaloid drugs, Sanguinarine (SG) and Berberine (BR), with biologically significant protonated and B-forms of herring sperm DNA in aqueous buffer medium of pH 3.5 and 7.4 respectively. The steady-state spectroscopic results substantiate the binding interaction of either drug molecule with protonated as well as B-form of DNA. The drug:DNA binding strength obtained employing the Benesi-Hildebrand equation reveals a significantly higher binding affinity of BR with B-DNA at pH 7.4 compared to that with protonated DNA at pH 3.5. However, the binding constant obtained for interaction of iminium form of SG with protonated DNA is found to be an order of magnitude higher compared to that in case of interaction with B-DNA. Moreover, the steady-state emission results validate the role of preferential electrostatic interaction between the cationic forms of the drugs (cationic BR and the iminium form of SG) and the negatively charged phosphate backbone of B-DNA under physiological conditions at pH 7.4 which has been further corroborated by monitoring the effect of a strong electrolyte on drug:DNA interaction under the various experimental conditions. The circular dichroic (CD) spectroscopic studies reveal the probable intercalative binding of the drug molecules within DNA at either pH. The time-resolved fluorescence decay measurements along with the time-resolved fluorescence anisotropy studies further substantiate the binding interaction of the drugs with protonated as well as B-form of herring sperm DNA.

Lighting Up DNA with the Environment-Sensitive Bright Adenine Analogue qAN4.

The fluorescent adenine analogue qAN4 was recently shown to possess promising photophysical properties, including a high brightness as a monomer. Here we report the synthesis of the phosphoramidite of qAN4 and its successful incorporation into DNA oligonucleotides using standard solid-phase synthesis. Circular dichroism and thermal melting studies indicate that the qAN4-modification has a stabilizing effect on the B-form of DNA. Moreover, qAN4 base-pairs selectively with thymine with mismatch penalties similar to those of mismatches of adenine. The low energy absorption band of qAN4 inside DNA has its peak around 358 nm and the emission in duplex DNA is partly quenched and blue-shifted (ca. 410 nm), compared to the monomeric form. The spectral properties of the fluorophore also show sensitivity to pH; a property that may find biological applications. Quantum yields in single-stranded DNA range from 1-29 % and in duplex DNA from 1-7 %. In combination with the absorptive properties, this gives an average brightness inside duplex DNA of 275 M-1 cm-1 , more than five times higher than the most used environment-sensitive fluorescent base analogue, 2-aminopurine. Finally, we show that qAN4 can be used to advantage as a donor for interbase FRET applications in combination with adenine analogue qAnitro as an acceptor.

Spectrophotometric and physicochemical studies on the interaction of a new platinum(IV) complex containing the drug pregabalin with calf thymus DNA

This study was designed to examine the interaction of a platinum(IV) complex containing the drug pregabalin with calf-thymus DNA (ct-DNA) under physiological conditions. The interaction between Pt(IV) complex and ct-DNA was investigated using ultraviolet-visible (UV-vis) spectroscopy, fluorescence spectroscopy, circular dichroism (CD), viscosity, molecular docking simulation and cell culture studies. Absorption spectra of Pt(IV) complex showed hypochromism. The calculated binding constant (Kb) obtained from UV-vis absorption study was 3.6 × 104 M−1. Furthermore, CD study demonstrated stability of the right-handed B-form of DNA in the presence of the Pt(IV) complex. In fluorimeteric studies, the binding mode of Pt(IV) complex with DNA was studied using MB as a fluorescence probe. The results showed that the complex is able to completely displace the MB intercalative binder into DNA, indicating intercalative binding. In addition, significant change on the viscosity of ct-DNA showed that the Pt(IV) complex binds to DNA by intercalative binding. Molecular docking revealed that the Pt(IV) complex not only binds to DNA by intercalative mode, but also van der Waals interactions or hydrogen bonds are the main forces in the binding process. Furthermore, the anticancer activity of Pt(IV) complex was observed with almost 100% decrease in cancer cell counts of MCF-7 breast cancer cell lines.

XNA Synthesis and Reverse Transcription by Engineered Thermophilic Polymerases

The B-family polymerases of hyperthermophilic archaea have proven an exceptional platform for engineering polymerases with extended substrate spectra, despite multiple mechanisms for detecting and avoiding incorporation of non-cognate substrates. These polymerases can efficiently synthesize and reverse-transcribe a number of xenonucleic acids (XNAs) that differ significantly from the canonical B-form of DNA. We present here a protocol for hexitol nucleic acid (HNA) synthesis by an engineered Thermococcus gorgonarius polymerase variant, including adaptation for large-scale synthesis and purification, and for other XNAs. We describe XNA purification and reverse transcription (with a previously reported XNA RT also based on Thermococcus gorgonarius), as well as key considerations for the characterization and optimization of XNA reactions. © 2018 by John Wiley & Sons, Inc.

DFT study of geometrical and vibrational features of a 3′,5′-deoxydisugar-monophosphate (dDSMP) DNA model in the presence of counterions and solvent

The B3LYP/6-31++G* theoretical level was used to study the influence of various hexahydrated monovalent (Li+, Na+, K+) and divalent (Mg2+) metal counterions in interaction with the charged PO2- group, on the geometrical and vibrational characteristics of the DNA fragments of 3',5'-dDSMP, represented by four conformers (g+g+, g+t, g-g- and g-t). All complexes were optimized through two solvation models [the explicit model (6H2O) and the hybrid model (6H2O/Continuum)]. The results obtained established that, in the hybrid model, counterions (Li+, Na+, K+, Mg2+) always remain in the bisector plane of the O1-P-O2 angle. When these counterions are explicitly hydrated, the smallest counterions (Li+, Na+) deviate from the bisector plane, while the largest counterions (K+ and Mg2+) always remain in the same plane. On the other hand, the present calculations reveal that the g+g+ conformer is the most stable in the presence of monovalent counterions, while conformers g+t and g-t are the most stable in the presence of the divalent counterion Mg2+. Finally, the hybrid solvation model seems to be in better agreement with the available crystallographic and spectroscopic (Raman) experiments than the explicit model. Indeed, the six conformational torsions of the C4'-C3'-O3'-PO-2-O5'-C5'-C4' segment of all complexes of the g-g- conformer in 6H2O/Continuum remain similar to the available experimental data of A- and B-DNA forms. The calculated wavenumbers of the g+g+ conformer in the presence of the monovalent counterion and of g-t conformer in presence of the divalent counterion in the hybrid model are in good agreement with the Raman experimental data of A- and B-DNA forms. In addition, the maximum deviation between the calculated wavenumbers in the 6H2O/Continuum for the g+g+ conformer and experimental value measured in an aqueous solution of the DMP-Na+ complex, is <1.07% for the PO2- (asymmetric and symmetric) stretching modes and <2.03% for the O5'-C5' and O3'-C3' stretching modes. Graphical abstract dDSMP-(OO)- Mg2+/6W/Continuum.

AMOEBA Polarizable Atomic Multipole Force Field for Nucleic Acids.

The AMOEBA polarizable atomic multipole force field for nucleic acids is presented. Valence and electrostatic parameters were determined from high-level quantum mechanical data, including structures, conformational energy, and electrostatic potentials, of nucleotide model compounds. Previously derived parameters for the phosphate group and nucleobases were incorporated. A total of over 35 μs of condensed-phase molecular dynamics simulations of DNA and RNA molecules in aqueous solution and crystal lattice were performed to validate and refine the force field. The solution and/or crystal structures of DNA B-form duplexes, RNA duplexes, and hairpins were captured with an average root-mean-squared deviation from NMR structures below or around 2.0 Å. Structural details, such as base pairing and stacking, sugar puckering, backbone and χ-torsion angles, groove geometries, and crystal packing interfaces, agreed well with NMR and/or X-ray. The interconversion between A- and B-form DNAs was observed in ethanol-water mixtures at 328 K. Crystal lattices of B- and Z-form DNA and A-form RNA were examined with simulations. For the RNA tetraloop, single strand tetramers, and HIV TAR with 29 residues, the simulated conformational states, 3 J-coupling, nuclear Overhauser effect, and residual dipolar coupling data were compared with NMR results. Starting from a totally unstacked/unfolding state, the rCAAU tetranucleotide was folded into A-form-like structures during ∼1 μs molecular dynamics simulations.

Melting Transition of Oriented DNA Fibers Submerged in Poly(ethylene glycol) Solutions Studied by Neutron Scattering and Calorimetry.

The influence of molecular confinement on the melting transition of oriented Na-DNA fibers submerged in poly(ethylene glycol) (PEG) solutions has been studied. The PEG solution exerts an osmotic pressure on the fibers which, in turn, is related to the DNA intermolecular distance. Calorimetry measurements show that the melting temperature increases and the width of the transition decreases with decreasing intermolecular distance. Neutron scattering was used to monitor the integrated intensity and width of a Bragg peak from the B-form of DNA as a function of temperature. The data were quantitatively analyzed using the Peyrard-Bishop-Dauxois model. The experiments and analysis showed that long segments of double-stranded DNA persist until the last stages of melting and that there appears to be a substantial increase of the DNA dynamics as the melting temperature of the DNA is approached.

FT-IR Spectroscopy Study in Early Diagnosis of Skin Cancer.

Mid-infrared spectroscopy (4000-500 cm-1) was used to analyze the spectral changes and differences of the characteristic absorption bands of the skin components due to cancer development for early clinical diagnosis. Human biopsies from basal cell carcinoma, malignant melanoma, and nevus were used, while normal skin tissue served as a control. The high quality of Fourier-transform infrared (FT-IR) spectra showed that upon cancer development the intensity of the absorption band at approximately 3062 cm-1 was increased, indicating that most of the proteins had the configuration of amide B and the β-sheet protein structure predominated. The stretching vibration bands of vCH2 in the region 2950-2850 cm-1 were increased in melanoma and nevus, while were less pronounced in basal cell carcinoma due to the increased lipophilic environment. In addition, the intensity of a new band at 1744 cm-1, which is assigned to aldehyde, was increased in melanoma and nevus and appeared as a shoulder in the spectra of normal skin. The absorption band of amide I at 1650 cm-1 was split into two bands, at 1650 cm-1 and 1633 cm-1, due to the presence of both α-helix and random coil protein conformations for melanoma and nevus. This was confirmed from the amide II band at 1550 cm-1, which shifted to lower frequencies at 1536 cm-1 and 1540 cm-1 for basal cell carcinoma and melanoma, respectively, indicating a damage of the native structure of proteins. The bands at 841 and 815 cm-1, which are assigned to B-DNA and Z-DNA, respectively, indicated that only the bands of the cancerous Z-DNA form are pronounced in melanoma, while in BCC both the characteristic bands of B-DNA and Z-DNA forms are found. It is proposed that the bands described above could be used as "diagnostic marker" bands for DNA forms, in the diagnosis of skin cancer.

Polarizable Force Field for DNA Based on the Classical Drude Oscillator: II. Microsecond Molecular Dynamics Simulations of Duplex DNA.

The structure and dynamics of DNA are governed by a sensitive balance between base stacking and pairing, hydration, and interactions with ions. Force-field models that include explicit representations of electronic polarization are capable of more accurately modeling the subtle details of these interactions versus commonly used additive force fields. In this work, we validate our recently refined polarizable force field for DNA based on the classical Drude oscillator model, in which electronic degrees of freedom are represented as negatively charged particles attached to their parent atoms via harmonic springs. The previous version of the force field, called Drude-2013, produced stable A- and B-DNA trajectories on the order of hundreds of nanoseconds, but deficiencies were identified that included weak base stacking ultimately leading to distortion of B-DNA duplexes and unstable Z-DNA. As a result of extensive refinement of base nonbonded terms and bonded parameters in the deoxyribofuranose sugar and phosphodiester backbone, we demonstrate that the new version of the Drude DNA force field is capable of simulating A- and B-forms of DNA on the microsecond time scale and the resulting conformational ensembles agree well with a broad set of experimental properties, including solution X-ray scattering profiles. In addition, simulations of Z-form duplex DNA in its crystal environment are stable on the order of 100 ns. The revised force field is to be called Drude-2017.

Non-enolisable Knoevenagel condensate appended Schiff bases-metal (II) complexes: Spectral characteristics, DNA-binding and nuclease activities

New Schiff base complexes [Cu(L1)Cl] (1), [Ni(L1)Cl] (2), [Zn(L1)Cl] (3), and [Fe(L2)H2OCl] (4) {L1=(4E)-3-(2-hydroxybenzylidene)-4-(2-hydroxyphenylimino)pentan-2-one, L2=2,2′-(1E,1′E)-(3-(2-hydroxybenzylidene)-pentane-2,4-diylidene)bis(azan-1-yl-1 idene)diphenol} have been synthesized and characterized by elemental analysis, UV–Vis, IR, FAB-mass, EPR, spectral studies and electrochemical studies, the ligands L1 &L2 were characterized by 1H and 13C NMR spectra. Complex 1 show a visible spectral d-d band near 600nm and display cyclic voltammetric quasireversible response for the Cu(II)/Cu(I) couple vs Ag/AgCl in DMSO. The EPR spectrum of 1 show g‖>g⊥ suggesting a square planar geometry around copper with dx2−y2 as the ground state. The mass spectral results have confirmed the proposed structure for complexes 1–4. DNA binding properties of these complexes 1–4 have been investigated by absorption titrations, cyclic voltammetric studies and circular dichroism studies. On titration with DNA, the complexes 1–4 show hypochromism at the MLCT band (13–31%) with a red shift of 1–8nm in the electronic spectrum and positive shift of voltammetric E1/2 in the CV studies are in favour of intercalative binding. CD spectra of 1 showed an increase in molar ellipticity (θ278) of the positive band with a minor red shift indicating the transition of B-form of DNA to A like form. DNA cleavage studies of complexes 1 and 4 with pUC18 DNA were studied by gel electrophoresis and complex 4 cleaves supercoiled pUC18 DNA in an oxidative manner in the presence of H2O2 and on photo irradiation at 312nm.

Synthesis and crystal structure elucidation of new copper(II)-based chemotherapeutic agent coupled with 1,2-DACH and orthovanillin: Validated by in vitro DNA/HSA binding profile and pBR322 cleavage pathway

New copper(II)-based complex (1) was synthesized and characterized by analytical, spectroscopic and single crystal X-ray diffraction. The in vitro binding studies of complex 1 with CT DNA and HSA have been investigated by employing biophysical techniques to examine the binding propensity of 1 towards DNA and HSA. The results showed that 1 avidly binds to CT DNA via electrostatic mode along with the hydrogen bonding interaction of NH2 and CN groups of Schiff base ligand with the base pairs of DNA helix, leads to partial unwinding and destabilization of the DNA double helix. Moreover, the CD spectral studies revealed that complex 1 binds through groove binding interaction that stabilizes the right-handed B-form of DNA. Complex 1 showed an impressive photoinduced nuclease activity generating single-strand breaks in comparison with the DNA cleavage activity in presence of visible light. The mechanistic investigation revealed the efficiency of 1 to cleave DNA strands by involving the generation of reactive oxygen species. Furthermore, the time dependent DNA cleavage activity showed that there was gradual increase in the amount of NC DNA on increasing the photoexposure time. However, the interaction of 1 and HSA showed that the change of intrinsic fluorescence intensity of HSA was induced by the microenvironment of Trp residue.

Studies of the B-Z transition of DNA: The temperature dependence of the free-energy difference, the composition of the counterion sheath in mixed salt, and the preparation of a sample of the 5'-d[T-(m(5) C-G)12 -T] duplex in pure B-DNA or Z-DNA form.

It is often envisioned that cations might coordinate at specific sites of nucleic acids and play an important structural role, for instance in the transition between B-DNA and Z-DNA. However, nucleic acid models explicitly devoid of specific sites may also exhibit features previously considered as evidence for specific binding. Such is the case of the "composite cylinder" (or CC) model which spreads out localized features of DNA structure and charge by cylindrical averaging, while sustaining the main difference between the B and Z structures, namely the better immersion of the B-DNA phosphodiester charges in the solution. Here, we analyze the non-electrostatic component of the free-energy difference between B-DNA and Z-DNA. We also compute the composition of the counterion sheath in a wide range of mixed-salt solutions and of temperatures: in contrast with the large difference of composition between the B-DNA and Z-DNA forms, the temperature dependence of sheath composition, previously unknown, is very weak. In order to validate the model, the mixed-salt predictions should be compared to experiment. We design a procedure for future measurements of the sheath composition based on Anomalous Small-Angle X-ray Scattering and complemented by (31) P NMR. With due consideration for the kinetics of the B-Z transition and for the capacity of generating at will the B or Z form in a single sample, the 5'-d[T-(m(5) C-G)12 -T] 26-mer emerges as a most suitable oligonucleotide for this study. Finally, the application of the finite element method to the resolution of the Poisson-Boltzmann equation is described in detail. © 2016 Wiley Periodicals, Inc. Biopolymers 105: 369-384, 2016.

Spectroscopic Studies of the Binding Interactions of Phenothiazininum Dyes (Thionine Acetate, Azure A and Azure B) with Calf-thymus DNA

The double helical structure of DNA offers various binding sites for the interaction of ligands or proteins. Interactions using minor groove, major groove, and through intercalation are the major types of binding mechanisms of DNA-ligand interactions. The lowering in the absorption intensity along with bathochromic shift is the indication of intercalation binding mode of the dye into the base pairs of the DNA. In this study, the interaction of phenothiazine dyes with calf-thymus DNA (ctDNA) in physiological buffer (pH 7.4) was studied using UV-visible, fluorescence, circular dichroism (CD), and UV-thermal denaturation spectroscopy. The binding constants were calculated at different temperatures with the help of fluorescence spectroscopy. CD signals signify that B-form of DNA might become more compact, upon binding of the dyes. Also, induced circular dichroism is observed which confirms the dye-DNA complex formation. Stabilization of DNA double helix upon binding with dyes was confirmed by the increase in Tm of ctDNA. Based on thermal melting profiles, it was found that thionine acetate is most promising in stabilizing the DNA double helix, in comparison to other two dyes. Also, binding constants calculated by fluorescence is in accordance with the thermal melting analysis. These results are indicative of the intercalation binding mode between dyes and the DNA. The binding affinity of the dyes to DNA is found to be in order as thionine acetate > azure A > azure B. Such preliminary studies facilitate our understanding about various types of DNAligand interactions and provide clues for designing new and more effective drugs.