Phase Behavior of Columnar DNA Assemblies

The pair interaction between two stiff parallel linear DNA molecules depends not only on the distance between their axes but on their azimuthal orientation. The positional and orientational order in columnar B-DNA assemblies in solution is investigated, based on the DNA-DNA electrostatic pair potential that takes into account DNA helical symmetry and the amount and distribution of adsorbed counterions. A phase diagram obtained by lattice sum calculations predicts a variety of positionally and azimuthally ordered phases and bundling transitions strongly depending on the counterion adsorption patterns.Comment: 4 pages, 3 figures, submitted to PR

Structural characterization of cationic lipid–tRNA complexes

Despite considerable interest and investigations on cationic lipid–DNA complexes, reports on lipid–RNA interaction are very limited. In contrast to lipid–DNA complexes where lipid binding induces partial B to A and B to C conformational changes, lipid–tRNA complexation preserves tRNA folded state. This study is the first attempt to investigate the binding of cationic lipid with transfer RNA and the effect of lipid complexation on tRNA aggregation and condensation. We examine the interaction of tRNA with cholesterol (Chol), 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP), dioctadecyldimethylammoniumbromide (DDAB) and dioleoylphosphatidylethanolamine (DOPE), at physiological condition, using constant tRNA concentration and various lipid contents. FTIR, UV-visible, CD spectroscopic methods and atomic force microscopy (AFM) were used to analyze lipid binding site, the binding constant and the effects of lipid interaction on tRNA stability, conformation and condensation. Structural analysis showed lipid–tRNA interactions with G–C and A–U base pairs as well as the backbone phosphate group with overall binding constants of KChol = 5.94 (± 0.8) × 104 M–1, KDDAB = 8.33 (± 0.90) × 105 M–1, KDOTAP = 1.05 (± 0.30) × 105 M–1 and KDOPE = 2.75 (± 0.50) × 104 M–1. The order of stability of lipid–tRNA complexation is DDAB > DOTAP > Chol > DOPE. Hydrophobic interactions between lipid aliphatic tails and tRNA were observed. RNA remains in A-family structure, while biopolymer aggregation and condensation occurred at high lipid concentrations

Structural characterization of cationic lipid–tRNA complexes

Despite considerable interest and investigations on cationic lipid–DNA complexes, reports on lipid–RNA interaction are very limited. In contrast to lipid–DNA complexes where lipid binding induces partial B to A and B to C conformational changes, lipid–tRNA complexation preserves tRNA folded state. This study is the first attempt to investigate the binding of cationic lipid with transfer RNA and the effect of lipid complexation on tRNA aggregation and condensation. We examine the interaction of tRNA with cholesterol (Chol), 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP), dioctadecyldimethylammoniumbromide (DDAB) and dioleoylphosphatidylethanolamine (DOPE), at physiological condition, using constant tRNA concentration and various lipid contents. FTIR, UV-visible, CD spectroscopic methods and atomic force microscopy (AFM) were used to analyze lipid binding site, the binding constant and the effects of lipid interaction on tRNA stability, conformation and condensation. Structural analysis showed lipid–tRNA interactions with G–C and A–U base pairs as well as the backbone phosphate group with overall binding constants of KChol = 5.94 (± 0.8) × 104 M–1, KDDAB = 8.33 (± 0.90) × 105 M–1, KDOTAP = 1.05 (± 0.30) × 105 M–1 and KDOPE = 2.75 (± 0.50) × 104 M–1. The order of stability of lipid–tRNA complexation is DDAB > DOTAP > Chol > DOPE. Hydrophobic interactions between lipid aliphatic tails and tRNA were observed. RNA remains in A-family structure, while biopolymer aggregation and condensation occurred at high lipid concentrations

Attraction between DNA molecules mediated by multivalent ions

The effective force between two parallel DNA molecules is calculated as a function of their mutual separation for different valencies of counter- and salt ions and different salt concentrations. Computer simulations of the primitive model are used and the shape of the DNA molecules is accurately modelled using different geometrical shapes. We find that multivalent ions induce a significant attraction between the DNA molecules whose strength can be tuned by the averaged valency of the ions. The physical origin of the attraction is traced back either to electrostatics or to entropic contributions. For multivalent counter- and monovalent salt ions, we find a salt-induced stabilization effect: the force is first attractive but gets repulsive for increasing salt concentration. Furthermore, we show that the multivalent-ion-induced attraction does not necessarily correlate with DNA overcharging.Comment: 51 pages and 13 figure

The Persistence Length of a Strongly Charged, Rod-like, Polyelectrolyte in the Presence of Salt

The persistence length of a single, intrinsically rigid polyelectrolyte chain, above the Manning condensation threshold is investigated theoretically in presence of added salt. Using a loop expansion method, the partition function is consistently calculated, taking into account corrections to mean-field theory. Within a mean-field approximation, the well-known results of Odijk, Skolnick and Fixman are reproduced. Beyond mean-field, it is found that density correlations between counterions and thermal fluctuations reduce the stiffness of the chain, indicating an effective attraction between monomers for highly charged chains and multivalent counterions. This attraction results in a possible mechanical instability (collapse), alluding to the phenomenon of DNA condensation. In addition, we find that more counterions condense on slightly bent conformations of the chain than predicted by the Manning model for the case of an infinite cylinder. Finally, our results are compared with previous models and experiments.Comment: 13 pages, 2 ps figure

Infrared Spectroscopic Studies of Cells and Tissues: Triple Helix Proteins as a Potential Biomarker for Tumors

In this work, the infrared (IR) spectra of living neural cells in suspension, native brain tissue, and native brain tumor tissue were investigated. Methods were developed to overcome the strong IR signal of liquid water so that the signal from the cellular biochemicals could be seen. Measurements could be performed during surgeries, within minutes after resection. Comparison between normal tissue, different cell lineages in suspension, and tumors allowed preliminary assignments of IR bands to be made. The most dramatic difference between tissues and cells was found to be in weaker IR absorbances usually assigned to the triple helix of collagens. Triple helix domains are common in larger structural proteins, and are typically found in the extracellular matrix (ECM) of tissues. An algorithm to correct offsets and calculate the band heights and positions of these bands was developed, so the variance between identical measurements could be assessed. The initial results indicate the triple helix signal is surprisingly consistent between different individuals, and is altered in tumor tissues. Taken together, these preliminary investigations indicate this triple helix signal may be a reliable biomarker for a tumor-like microenvironment. Thus, this signal has potential to aid in the intra-operational delineation of brain tumor borders. © 2013 Stelling et al

Adsorption of mono- and multivalent cat- and anions on DNA molecules

Adsorption of monovalent and multivalent cat- and anions on a deoxyribose nucleic acid (DNA) molecule from a salt solution is investigated by computer simulation. The ions are modelled as charged hard spheres, the DNA molecule as a point charge pattern following the double-helical phosphate strands. The geometrical shape of the DNA molecules is modelled on different levels ranging from a simple cylindrical shape to structured models which include the major and minor grooves between the phosphate strands. The densities of the ions adsorbed on the phosphate strands, in the major and in the minor grooves are calculated. First, we find that the adsorption pattern on the DNA surface depends strongly on its geometrical shape: counterions adsorb preferentially along the phosphate strands for a cylindrical model shape, but in the minor groove for a geometrically structured model. Second, we find that an addition of monovalent salt ions results in an increase of the charge density in the minor groove while the total charge density of ions adsorbed in the major groove stays unchanged. The adsorbed ion densities are highly structured along the minor groove while they are almost smeared along the major groove. Furthermore, for a fixed amount of added salt, the major groove cationic charge is independent on the counterion valency. For increasing salt concentration the major groove is neutralized while the total charge adsorbed in the minor groove is constant. DNA overcharging is detected for multivalent salt. Simulations for a larger ion radii, which mimic the effect of the ion hydration, indicate an increased adsorbtion of cations in the major groove.Comment: 34 pages with 14 figure

Interaction of Pyrrolobenzodiazepine (PBD) Ligands with Parallel Intermolecular G-Quadruplex Complex Using Spectroscopy and ESI-MS

Studies on ligand interaction with quadruplex DNA, and their role in stabilizing the complex at concentration prevailing under physiological condition, has attained high interest. Electrospray ionization mass spectrometry (ESI-MS) and spectroscopic studies in solution were used to evaluate the interaction of PBD and TMPyP4 ligands, stoichiometry and selectivity to G-quadruplex DNA. Two synthetic ligands from PBD family, namely pyrene-linked pyrrolo[2,1-c][1,4]benzodiazepine hybrid (PBD1), mixed imine-amide pyrrolobenzodiazepine dimer (PBD2) and 5,10,15,20-tetrakis(N-methyl-4-pyridyl)porphyrin (TMPyP4) were studied. G-rich single-stranded oligonucleotide d(5′GGGGTTGGGG3′) designated as d(T2G8), from the telomeric region of Tetrahymena Glaucoma, was considered for the interaction with ligands. ESI-MS and spectroscopic methods viz., circular dichroism (CD), UV-Visible, and fluorescence were employed to investigate the G-quadruplex structures formed by d(T2G8) sequence and its interaction with PBD and TMPyP4 ligands. From ESI-MS spectra, it is evident that the majority of quadruplexes exist as d(T2G8)2 and d(T2G8)4 forms possessing two to ten cations in the centre, thereby stabilizing the complex. CD band of PBD1 and PBD2 showed hypo and hyperchromicity, on interaction with quadruplex DNA, indicating unfolding and stabilization of quadruplex DNA complex, respectively. UV-Visible and fluorescence experiments suggest that PBD1 bind externally where as PBD2 intercalate moderately and bind externally to G-quadruplex DNA. Further, melting experiments using SYBR Green indicate that PBD1 unfolds and PBD2 stabilizes the G-quadruplex complex. ITC experiments using d(T2G8) quadruplex with PBD ligands reveal that PBD1 and PBD2 prefer external/loop binding and external/intercalative binding to quadruplex DNA, respectively. From experimental results it is clear that the interaction of PBD2 and TMPyP4 impart higher stability to the quadruplex complex

Some Schiff base complexes of the lanthanide and calcium ions.

Many new complexes of tri- and tetra- valent lanthanide ions and divalent calcium with Schiff bases have been synthesised and their properties investigated by chemical analysis, infra-red, visible, and ultra-violet absorption spectroscopy, X-ray powder diffraction studies, and molar conductance and magnetic susceptibility measurements. The Schiff bases, which were mainly derivatives of sali-cylaldehyde and primary amines, formed complexes in two ways. In the first, the phenolic hydrogen atoms were un -ionised and the base behaved as a neutral ligand whilst in the second, the base reacted in the usual way as an anion. For the trivalent metal ions, complexes of the following types were obtained:- 1. Ln(L)Cl[3]. nH[2]O and La(Hsal-ane)Cl[3]. [2]EtOH. (Ln=Ce,La; n=1 and 2; L=NN'-ethylenebis(salicylideneimine) (H[2]salen) and NN'-1,3-propylenebis(salicylideneimine) (H[2]sal-1,3pn); N-salicylideneaniline (Hsal-ane); EtOH-ethanol). 2. Ln[2](L)[3]X[6]. nH[2]O. 3. Ln(L)[2]X[3]. nH[2]O and La(Hsal-ane)[2](NO[3])[3]. 2EtOH. 4. Ln(L)[3]X[3]. nH[2]O, (Ln=La,Ce,Pr,Nd,Sm,Gd,Ho,Yb; X=Cl-,Br-,NO[3-]; L=H[2]salen, H[2]sal-1,3pn, NN'-1,2-propylenebis(salicylideneimine) (H2sal-1,2pn), NN'-1,6-hexylenebis(salicylideneimine) (H[2]sal-1,6hex), and Hsal-ane ; n=0 - 2). A few exceptions are discussed in detail in Chapter 2. New series of complexes with tetravalent cerium with ionised and un-ionised Schiff bases of formulae below have been synthesised:- 5. Ce(L')2, 6. Ce(L)2(NO3)4. H2O (L' = the ionised Schiff bases: NN' -1,2-propylenebis(salicylideneiminate) (sal-1,2pn), NN'-1,3-propylenebis(salicylideneiminate) (sal-1,3pn), NN'-ethylenebis(3-inethoxysalicylideneiminate) (3-methoxy-salen), NN'-ethylenebis(5-nitrosalicylideneiminate) (5-nitrosalen), NN'-1,2-propylenebis(5-nitrosalicylideneiminate) (5-nitrosal-1,2pn), NN'-1,3-propylenebis(5-nitrosalicylideneiminate) (5-nitrosal-1,3pn) and L=H2salen, H2sal-1,2pn, H2sal-1,3pn and H2sal-1,6hex). Calcium complexes of the types shown below have been isolated:- 7. Ca(L)X2. nsolvent, 8. Ca(L)2X2. nsolvent (n=0 - 2; solvent = H2O or ethanol; L=H2salen, H2sal-1,2pn, H2sal-1,3pn, and H2sal-1,6hex and X=Cl- and NO3-). On the basis of the infra-red spectroscopy it was concluded that the un-ionised ligands behave as bidentate species with coordination taking place through the azomethine nitrogen atoms and not through the phenolic oxygen atoms. The ionised bases behave as tetra-dentate ligands in the well-established manner. There is evidence for coordinated nitrate in all the lanthanide nitrato- complexes. Far infra-red spectroscopy has shown the presence of the coordinated halide in some of the lanthanide complexes. The coordination numbers of a series of neodymium(III) Schiff base complexes have been deduced through the hypersensitive transitions observed in the visible region of the absorption spectrum by comparison with the spectra of complexes of known structure. The coordination numbers of the lanthanide complexes are probably 8 or 9 in the halide complexes and between 10 and 12 for the nitrates. The calcium complexes may have coordination numbers between 4 and 6. Furthermore, the X-ray powder photography has shown the complexes to be true compounds and not simple mixtures. Isomorphism was found in some cases for the trivalent lanthanum and neodymium complexes. The conclusions were consistent with those obtained from the other measurements