Left-handed Double-helical Structure Research Articles

Revealing the nature of localized and delocalized excitations of DNA

Unmatched opportunities for novel nanoscale technologies and applications are provided by deoxyribonucleic acid (DNA). In this paper, we propose a quantum mechanical investigation into the optical excitations of DNA using calculations based on time-dependent density functional theory. Our findings demonstrate that DNA exhibits photoabsorption in the ultraviolet region of the electromagnetic spectrum, and transition contribution maps demonstrate that the optical transitions of DNA contain both localized and delocalized excitations. Additionally, we simulated and contrasted the electronic circular dichroism (ECD) spectra of B-DNA (a right-handed double helical structure) and Z-DNA (a left-handed double helical structure). We discovered significant variations in the ECD spectra. The design of DNA-based nano-optical devices should benefit from the theoretical findings in this work.

Linking Temperature, Cation Concentration and Water Activity for the B to Z Conformational Transition in DNA.

High concentrations of Na+ or [Co(NH3)6]3+ can induce the B to Z conformational transition in alternating (dC-dG) oligo and polynucleotides. The use of short DNA oligomers (dC-dG)4 and (dm5C-dG)4 as models can allow a thermodynamic characterization of the transition. Both form right handed double helical structures (B-DNA) in standard phosphate buffer with 115 mM Na+ at 25 °C. However, at 2.0 M Na+ or 200 μM [Co(NH3)6]3+, (dm5C-dG)4 assumes a left handed double helical structure (Z-DNA) while the unmethylated (dC-dG)4 analogue remains right handed under those conditions. We have previously demonstrated that the enthalpy of the transition at 25 °C for either inducer can be determined using isothermal titration calorimetry (ITC). Here, ITC is used to investigate the linkages between temperature, water activity and DNA conformation. We found that the determined enthalpy for each titration varied linearly with temperature allowing determination of the heat capacity change (ΔCp) between the initial and final states. As expected, the ΔCp values were dependent upon the cation (i.e., Na+ vs. [Co(NH3)6]3+) as well as the sequence of the DNA oligomer (i.e., methylated vs. unmethylated). Osmotic stress experiments were carried out to determine the gain or loss of water by the oligomer induced by the titration. The results are discussed in terms of solvent accessible surface areas, electrostatic interactions and the role of water.

Helical Structure and Circular Dichroism Spectra of DNA: A Theoretical Study

The helical structure is experimentally determined by circular dichroism (CD) spectra. The sign and shape of the CD spectra are different between B-DNA with a right-handed double-helical structure and Z-DNA with a left-handed double-helical structure. In particular, the sign at around 295 nm in CD spectra is positive for B-DNA, which is opposite to that of Z-DNA. However, it is difficult to determine the helical structure from the UV absorption spectra. Three important factors that affect the CD spectra of DNA are (1) the conformation of dG monomer, (2) the hydrogen-bonding interaction between two helices, and (3) the stacking interaction between nucleic acid bases. We calculated the CD spectra of (1) the dG monomer at different conformations, (2) the composite of dG and dC monomers, (3) two dimer models that simulate separately the hydrogen-bonding interaction and the stacking interaction, and (4) the tetramer model that includes both hydrogen-bonding and stacking interactions simultaneously. The helical structure of DNA can be clarified by a comparison of the experimental and SAC-CI theoretical CD spectra of DNA and that the sign at around 295 nm of the CD spectra of Z-DNA reflects from the strong stacking interaction characteristic of its helical structure.

Enthalpy of the B-to-Z Conformational Transition of a DNA Oligonucleotide Determined by Isothermal Titration Calorimetry

The influence of high concentrations of Na + or [Co(NH 3) 6] 3+ on the conformation of two related DNA oligomers was investigated by circular dichroism spectropolarimetry (CD), isothermal titration calorimetry (ITC), and differential scanning calorimetry (DSC). As revealed by CD, DNA oligomers, (dC-dG) 4 and (dm 5C-dG) 4, both form right-handed double helical structures (B-DNA) in standard phosphate buffer with 115 mM Na + at 25°C. However, at 2.0 M Na + or 200 μM [Co(NH 3) 6] 3+, (dm 5C-dG) 4 assumes a left-handed double helical structure (Z-DNA), whereas the unmethylated (dC-dG) 4 analog remains right-handed under those conditions. ITC was then used to determine the enthalpy change upon increasing the concentration of either Na + or [Co(NH 3) 6] 3+ for both DNA oligomers at 25°C. The titration with Na + resulted in endothermic isotherms with (dm 5C-dG) 4 being more endothermic than (dC-dG) 4 by 700 cal/mol basepair. In contrast, titration with [Co(NH 3) 6] 3+ resulted in exothermic isotherms with (dC-dG) 4 being more exothermic than (dm 5C-dG) 4 by 720 cal/mol basepair. We attribute the enthalpy difference to the conformational transition from B-form DNA to Z-form DNA for (dm 5C-dG) 4, a transition which does not occur for the unmethylated (dC-dG) 4. The value of ∼700 cal/mol basepair for the enthalpy of the B-Z transition compares favorably with previously published results obtained by different techniques. DSC was used to monitor the duplex to single strand transitions for both oligomers under the different concentrations. These results indicated that methylation of the cytidine destabilizes (dm 5C-dG) 4 relative to (dC-dG) 4. Coupling the DSC data with the ITC data allowed construction of a thermodynamic cycle which gives insight into the influence of both temperature and ionic strength on the heat content of the two DNA systems studied. Further, this study reveals the utility of using ITC for determinations of transition enthalpies with the appropriate choice of control.

Dianion-controlled supramolecular assembly of copper(II)-arginine complex ion

In order to clarify the self-organization and structure-determining factors in the [Cu(l- or d-Arg)2]2+-containing systems (Arg=arginine), we studied the solution and crystal structures of [Cu(l-Arg)2]2+-X2− systems, where X refers to dinegative tetrahedral anions, sulfate or 1,3-benzenedisulfonate (m-bs2−), by electronic absorption, circular dichroism (CD), and ESR spectroscopic methods and X-ray structural analysis. The molecular structure of [Cu(l-Arg)2](SO4) exhibited a planar Cu(II) geometry with the two coordinated amino groups in the cis configuration due to the hydrogen bonds with SO42−, and the complex with X2−=m-bs2− assumed the trans configuration without hydrogen bonds with m-bs2−. The crystal structure of [Cu(l-Arg)2](SO4) showed that the complex cations were bound with the anions to form a right-handed double-strand single-helical structure, whereas that of [Cu(l-Arg)2](m-bs2−) revealed that the complex ions were bound with each other through the guanidinium–coordinated carboxylate hydrogen bonds to form a 3-dimensional layer structure with the m-bs2− ions connecting the layers by hydrogen bonds. The d-Arg analog of the complex with isophthalate (m-pa2−) used in place of m-bs2−, [Cu(d-Arg)2](m-pa2−), was shown to assume the cis configuration and form a left-handed double-helical structure. The configuration of the Cu(II) plane was predictable on the basis of the structural requirements for the hydrogen bonding with X2−. Comparison of the absorption spectra of [Cu(l-Arg)2](X) in the solid state and in aqueous solution indicated that the complexes in solution are more in the cis configuration than in the trans. The CD spectra of the sulfate system exhibited a unique spectral behavior, which suggested formation of stronger hydrogen bonds between coordinated amino groups and the sulfate ion. Factors determining the observed self-organized structures were comparatively discussed in connection with unique infinite right- or left-handed double-helical structures of the [Cu(l- or d-Arg)2]2+–X2− systems reported previously (N. Ohata, H. Masuda, and O. Yamauchi, Angew. Chem., Int. Ed. Engl. 35 (1996) 531).

Molecular and crystal structure of d(CGCGmo 4CG): N4-methoxycytosine · guanine base-pairs in Z-DNA

The base analogue N 4-methoxycytosine (mo 4C) is ambivalent in its hydrogen-bonding potential, since it forms stable base-pairs with both adenine and guanine in oligomer duplexes. To investigate the base-pair geometry, the structure of d(CGCGmo 4CG) has been determined by single-crystal X-ray diffraction techniques. The d(CGCGmo 4CG) 2 crystallized in a left-handed double helical structure ( Z-type). Refinement using 2559 reflections between 10 and 1.7 Å converged with a final R = 0.181 ( Rw = 0.130) including 68 solvent molecules. The orthorhombic crystals are in the space group P2 12 12 1, with cell dimensions a= 18.17 A ̊ , b = 30.36 A ̊ , c = 43.93 A ̊ . The mo 4C · G base-pair is of the wobble type, with mo 4C in the imino form, and the methoxy group in the syn configuration.

Hybrid hexanucleotide duplex containing cyclonucleotides and deoxynucleotides: the d(TA) segment can adopt a high anti left-handed double-helical structure

It is known that oligonucleotides containing cyclonucleosides with a high anti (intermediate between anti and syn) glycosidic conformation adopt left-handed, single- and double-helical structures [Uesugi, S., Yano, J., Yano, E., & Ikehara, M. (1977) J. Am. Chem. Soc. 99, 2313-2323]. In order to see whether DNA can adopt the high anti left-handed double-helical structure or not, a self-complementary hexanucleotide containing 6,2'-O-cyclocytidine (C(o)), 8,2'-O-cycloguanosine (G(o)), thymidine, and deoxyadenosine, C(o)G(o)dTdAC(o)G(o), was synthesized. Imino proton NMR spectra and the results of nuclear Overhauser effect experiments strongly suggest that C(o)G(o)dTdAC(o)G(o) adopts a left-handed double-helical structure where the deoxynucleoside residues are involved in hydrogen bonding and take a high anti glycosidic conformation. A conformational model of the left-handed duplex was obtained by calculation with energy minimization. Thus it appears that DNA can form a high anti, left-handed double helix under some constrained conditions, which is quite different from that of Z-DNA.

Hybrid oligomer of cyclonucleotides and deoxynucleotides. A high anti left-handed double helical DNA structure.

It has been shown by us that oligonucleotides containing cyclonucleosides with a high anti glycosidic conformation take left-handed, single and double helical structures (S. Uesugi, J. Yano, E. Yano and M. Ikehara, J. Am. Chem. Soc. 99,2313 (1977) and references therein). In order to see whether DNA can adopt the high anti left-handed double helical structure or not, a self-complementary hexanucleotide containing 6,2'-O-cyclocytidine (C 0). 8,2'-O-cycloguanosine (G 0), deoxycytidine and deoxyguanosine, C 0 G 0 dCdGC 0 G 0, was synthesized. Corresponding hexanucleotide containing only cyclonucleosides, C 0 G 0 C 0 G 0 C 0 G 0, was also synthesized. Their conformation was examined by UV, CD and 1H NMR spectroscopy. C 0 G 0 C 0 G 0 C 0 G 0 forms an unusually stable, left-handed duplex. Imino proton NMR spectra and the results of nuclear Overhauser effect experiments strongly suggest that C 0 G 0 dCdGC0 G 0 take a left-handed double helical structure where the deoxynucleoside residues are involved in hydrogen bonding and take a high anti glycosidic conformation. Thus it is revealed that DNA could form a high anti, left-handed double helix which is different from that of Z-DNA under some constrained conditions.

Left handed DNA in synthetic and topologically constrained form V DNA and its implications in protein recognition

We have investigated structural transitions in Poly(dG-dC) and Poly(dG-Me5dC) in order to understand the exact role of cations in stabilizing left-handed helical structures in specific sequences andthe biological role, if any, of these structures. From a novel temperature dependent Z ⇌ B transition it has been shown that a minor fluctuation in Na+ concentration at ambient temperature can bring about B to Z transition. Forthe first time, wehave observed a novel Z⇌B⇌Zuble transition in poly(dG-Me5dC) as the Na+ concentration is gradually increased. This suggests that a minor fluctuation in Na+ concentration in conjunction with methylation may transform small stretches of CG sequences from one conformational state to another. These stretches could probably serve as sites for regulation. Supercoiled formV DNA reconstituted from pBR322 and pβG plasmids have been studied as model systems, in order to understand the nature and role of left-handed helical conformation in natural sequences. A large portion of DNA in form V, obtained by reannealing the two complementary singlestranded circles is forced to adopt left-handed double helical structure due to topological constraints (Lk = 0). Binding studies with Z-DNA specific antibody and spectroscopic studies confirm the presence of left-handed Z-structure in the pβG and pβR322 form V DNA. Cobalt hexamine chloride, which induces Z-form in Poly(dG-dC) stabilizes the Z-conformation in form V DNA even in the non-alternating purine-pyrimidine sequences. A reverse effect is observed with ethidium bromide. Interestingly, both topoisomerase I and II (from wheat germ) act effectively on form V DNA to give rise to a species having an electrophoretic mobility on agarose gel similar to that of open circular (form II) DNA. Whether this molecule is formed as a result of the left-handed helical segments of form V DNA undergoing a transition to the right-handed B-form during the topoisomerase action remains to be solved.

Salt-induced transition between two double-helical forms of oligo (dC-dG).

The change between a right-handed and a left-handed double-helical structure is certainly an extreme example of the conformational variability of DNA. Studying the energetics and dynamics with defined oligomers provides a convenient way to obtain a quantitative understanding of such processes and can serve as a model for possible regulatory mechanisms in biological systems. Monoclonal antibodies with different specificities to Z-DNA offer a promising technique to search for such structures in more-complicated systems and represent a model system for the quantitative investigation of the interaction of proteins with particular conformations of DNA.

Z-DNA: vacuum ultraviolet circular dichroism.

In concentrated salt or ethanolic solutions, the self-complementary copolymer poly(dG-dC).poly(dG-dC) forms a left-handed double-helical structure that has been termed "Z-DNA." The first evidence for this structure came from changes observed in the circular dichroism (CD) spectrum between 230 and 300 nm for low- and high-salt solutions (Pohl, F. M. & Jovin, T. M. (1972) J. Mol. Biol. 67, 675-696). In 3 M NaCl, the CD spectrum is approximately inverted compared to the B-form spectrum observed in low-salt solution. We measured the vacuum ultraviolet CD spectrum of poly(dG-dC).poly(dG-dC) down to 180 nm under conditions in which the 230- to 300-nm spectrum is inverted. Below 200 nm, where the B form exhibits the large positive peak at 187 nm that is characteristic of right-handed double-helical DNAs, the Z form exhibits a large negative peak at 194 nm and a positive band below 186 nm. Therefore, the Z-form vacuum ultraviolet CD spectrum resembles an inverted and red-shifted B-form spectrum. The magnitudes of the differences observed between the B and Z forms in the CD spectrum below 200 nm are about 10 times greater than those observed between 230 and 300 nm. The vacuum ultraviolet CD spectrum of poly(dG-dC).poly(dG-dC) in 3 M Cs2SO4 also is inverted compared to the B-form spectrum; however, between 230 and 300 nm, it is nonconservative with a negative maximum at 290 nm and a weak positive CD signal above 300 nm, presumably reflecting differential light scattering and indicating the existence of molecular aggregates. Our results suggest that the vacuum ultraviolet CD spectrum is sensitive to the handedness of double-helical DNA structures. The CD spectrum in this region should complement other spectroscopic methods in relating the structures of poly(dG-dC).poly(dG-dC) existing in solution to those determined in the solid state by x-ray crystallography.

Induced cotton effects of hyaluronic acid-acridine orange complex and conformation of the polymer

Hyaluronic acid-acridine orange complexes in water show a symmetric doublet CD band centered at 455 nm. The exciton-like CD band indicates a preferred left-handed chirality for this polymer in solution, which correlates with the proposed left-handed double helical structure of hyaluronic acid in oriented film. The sign and general shape of the induced CD band with hyaluronic acid (dye concentration 2.5×10 −5M) is the same throughout the range of polymer-dye ratio studied. The single CD band which appears at a 10 −4M concentration of dye has been attributed to dye-dye interaction. Hyaluronic acid, preheated to 95°C for one hour, followed by rapid cooling, does not show any induced CD band.