Proton relaxation in aqueous DNA solutions

By use of D₂O we found that the shortening of the longitudinal proton relaxation time which occurs in the investigated aqueous yeast DNA solutions (≦ 2.4% with 2% protein) was not based on a hydration effect, but was caused by magnetic impurities only. An estimate shows that the mobility of the hydrated water molecules is reduced by less than two orders of magnitude in comparison with the free water molecules.     

Properties of monovalent nickel complexes with tetraaza-macrocyclic ligands in aqueous solutions

The effects of N-alkylation on the redox potential of the couples NiLⁱ²⁺/NiLⁱ⁺, L = tetraaza-14-membered-macrocyclic ligands, and on the properties of the monovalent nickel complexes in aqueous solutions are reported for 14 complexes. The spectra and lifetimes of the NiLⁱ⁺ complexes are reported. The self-exchange rates for the couples NiLⁱ²⁺/NiLⁱ⁺ were determined. Two of the ligands were synthesized for the first time for this study. Cyclic voltammetry and pulse radiolysis were used. The results point out that: (i) N-alkylation always shifts the redox potential to a less cathodic one; this effect stems to a large degree from the decrease in the solvation energy of the complex caused by the N-alkylation of the ligand. (ii). The lifetime of the monovalent complexes is not linearly related to the redox potential of the NiLⁱ²⁺/NiLⁱ⁺ couples. (iii) The NiLⁱ⁺ complexes exist in several isomeric forms; the rate of the isomerization depends on the structure of the ligand. (iv) Different isomers of NiLⁱ⁺ may be formed when the complex NiLⁱ²⁺ is reduced by different reagents; therefore, the pulse-radiolytically formed NiLⁱ⁺ complexes might have different properties than those formed electrochemically.     

DNA conformation and thermostability in urea aqueous solutions

It is suggested from the character of the change of circular dichroism spectra that in the presence of urea winding of DNA double helix takes place within the bounds of B-family of forms. It is shown that the realized conformation of DNA differs from the experimentally known forms of DNA belonging to B-family. Urea destabilizes the DNA molecules without connection with helical and melt pairs of DNA nitrous bases. Urea affects the conformational state of DNA by water destruction around DNA, which is accompanied by dehydration of DNA and basic metal ions.     

Formation of hydrogen peroxide by VUV-photolysis of water and aqueous solutions with methanol

The hydrogen peroxide production upon vacuum ultraviolet (VUV) irradiation of water is reviewed, because published results from the last 10 years lead to conflicting mechanistic interpretations. This work confirms that in pure water, hydrogen peroxide is only produced in the presence of molecular oxygen. Mechanistic schemes explain these findings and confirm earlier statements that recombination of hydroxyl radicals is kinetically disfavoured. In agreement with other recent publications, this work confirms that enhanced hydrogen peroxide production takes place upon VUV irradiation of aqueous solutions of organic compounds. For these investigations, methanol was chosen as an organic model compound. During photolyses, hydrogen peroxide, dissolved molecular oxygen, pH-value of the reaction system, methanol and its products of oxidative degradation were analyzed, and kinetic studies were undertaken to explain the evolution of the concentrations of these components.     

Stability and thermodynamics of polyuridylic acid-deoxyadenosine complexes in aqueous neutral salt solutions

Using the thermodynamic analysis and methodology of Hill (Biopolymers, 12 , 257 (1973)) for the treatment of optical thermal transition data the effects of various neutral salt additives on the stability and thermodynamics of the poly U–deoxyadenosine interactions that lead to the formation of triple-stranded helical polymer–monomer complexes have been studied. In order of increasing molar effectiveness as polyU–deoxyadenosine complex stability perturbants (pH 7 and in the presence of 1 M NaCl), the various ions may be ranked: SO₄^?2 < Cl^? < Br^? < ClO₄^?; and (CH₃)₄ N⁺ < Li⁺ < Rb⁺ ～ Na⁺ < K⁺ < (CH₃ CH₂)₄ N⁺ < urea < Guan⁺ ～ (CH₃(CH₂)₂)₄ N⁺. Destabilizing neutral salt additives (e.g., NaClO₄) caused a decrease in the absolute magnitude of the apparent enthalpy and entropy of binding relative to the values determined in the presence of NaCl. By contrast, stabilizing additives (e.g., Na₂ SO₄) had the opposite effect on these parameters. Along a melting curve the apparent differential heat of complex formation calculated for the binding of deoxyadenosine to poly U in 1 M NaCl appeared to vary linearly with θ, the extent of fractional binding. For such a linear dependence it can be shown that the integral heat (usually determined calorimetrically) equals the differential heat at θ = 0.5. Correcting the apparent differential heat calculated at θ = 0.5 for ligand activity resulted in values for the integral heat of binding of deoxyadenosine to poly U in 1 M NaCl of ?13 to ?16 kcal/mol. Binding isotherms determined in the presence of different inorganic electrolytes could be superimposed provided that different temperatures were compared. However, the additive (CH₃)₄NCl, which has been shown to interact preferentially with A-T rich regions of DNA (Shapiro, Stannard, and Felsenfeld, Biochemistry, 8 , 3233 (1969)) resulted in a considerably broadened binding isotherm indicating less cooperativity.     

Atomic force microscopy of DNA in aqueous solutions.

DNA on mica can be imaged in the atomic force microscope (AFM) in water or in some buffers if the sample has first been dehydrated thoroughly with propanol or by baking in vacuum and if the sample is imaged with a tip that has been deposited in the scanning electron microscope (SEM). Without adequate dehydration or with an unmodified tip, the DNA is scraped off the substrate by AFM-imaging in aqueous solutions. The measured heights and widths of DNA are larger in aqueous solutions than in propanol. The measured lengths of DNA molecules are the same in propanol and in aqueous solutions and correspond to the base spacing for B-DNA, the hydrated form of DNA; when the DNA is again imaged in propanol after buffer, however, it shortens to the length expected for dehydrated A-DNA. Other results include the imaging of E. coli RNA polymerase bound to DNA in a propanol-water mixture and the observation that washing samples in the AFM is an effective way of disaggregating salt-DNA complexes. The ability to image DNA in aqueous solutions has potential applications for observing processes involving DNA in the AFM.     

Acidity and photolability of ruthenium salen nitrosyl and aquo complexes in aqueous solutions

The photochemical behavior of nitrosyl complexes Ru(salen)(NO)(OH₂)⁺ and Ru(salen)(NO)Cl (salen = N,N′-ethylenebis-(salicylideneiminato) dianion) in aqueous solution is described. Irradiation with light in the 350-450 nm range resulted in nitric oxide (NO) release from both. For Ru(salen)(NO)Cl secondary photoreactions also resulted in chloride aquation. Thus, in both cases the final photoproduct is the diaquo cation , for which pK_a’s of 5.9 and 9.1 were determined for the coordinated waters. The pK_a of the Ru(salen)(NO)(OH₂)⁺ cation was also determined as 4.5 ± 0.1, and the relative acidities of these ruthenium aquo units are discussed in the context of the bonding interactions between Ru(III) and NO.     

Competitive reactions in solutions of DNA and water-soluble interpolyelectrolyte complexes

The reaction of competitive binding of two polyanions—DNA and synthetic fluorescence-tagged poly(methacrylate) (PMA*)—with the polycation-quencher poly(N-ethyl-4-vinyl-pyridinium) (PEVP) was studied by fluorescence quenching technique. It was found that ability of DNA to displace PMA *from the water-soluble nonstoichiometric interpolyelectrolyte complex (NPEC) formed by PMA* and PEVP—NPEC(PMA*-PEVP)—and to form water-soluble NPEC(DNA-PEVP) can be determined by the parameter Ψ = P _PMA*/P _PEVP where P _PMA* and P _PEVP are the degrees of polymerization of PMA* and PEVP, respectively. In the case of Ψ < 1 the decrease of Ψ leads to the shift of the reaction equilibrium to the right, which can be explained by the gain of entropy due to the increase of the total number of polymeric particles in the solution. Introduction of alkali metal cations into the reaction mixture results in the shift of the reaction equilibrium, and according to their ability to shift the equilibrium to the right the cations can be arranged in the series Na⁺ > K⁺ > Li⁺. The substitution of native DNA by denatured DNA practically does not affect the reaction equilibrium in solutions of NaCl and KCl but considerably shifts it to the right in solutions of LiCl. The data obtained are in accordance with the differences in the selectivity of alkali cations binding with competitive polyanions. © 1995 John Wiley & Sons, Inc.     

Vibrational circular dichroism of carbohydrate films formed from aqueous solutions

Vibrational circular dichroism (VCD) spectra in the entire 2000-900 cm(-1) region have been recorded, for the first time, for films of carbohydrates prepared from aqueous solutions. Eight different carbohydrates, alpha-D-glucopyranosyl-(1-->4)-D-glucose, cyclomaltohexaose, alpha-D-glucopyranosyl alpha-D-glucopyranoside, beta-D-glucopyranosyl-(1-->6)-D-glucose, beta-D-glucopyranosyl-(1-->4)-D-glucose, D-glucose, and both enantiomers of 6-deoxygalactose and of allose, were investigated. The VCD spectra obtained for films are found to be identical to the corresponding spectra obtained for aqueous solutions of carbohydrates. These measurements demonstrate several advantages of significant importance. The strong infrared absorption of water has prevented, in the past, the pursuit for routine applications of VCD in determining the structures of carbohydrates in aqueous solutions. This limitation is not present for film studies because water solvent is removed in the process of preparing the films. Also, strong infrared absorption of water at 1650 cm(-1) requires the use of very short-pathlength (6 microm) cells for measurements on aqueous solutions. This requirement and concomitant inconveniences (such as laborious assembling of a demountable liquid cell or purchasing an expensive variable pathlength liquid cell) have been eliminated for film measurements. The removal of interfering water absorption in film studies resulted in higher light throughput and better signal-to-noise ratios for VCD measurements. Another point of significance is that the amount of carbohydrate sample required for VCD measurements on films is approximately one to two orders of magnitude smaller than that required for corresponding VCD measurements on aqueous solutions. Since carbohydrate samples can now be studied as films, VCD spectroscopy becomes much more broadly applicable for carbohydrates than previously believed. The present work, in combination with other film measurements in our laboratory, indicate that VCD studies on films can be used more generally, providing a convenient and powerful approach for probing structural information for biologically important compounds.     

Formation and structural determinants of multi-stranded guanine-rich DNA complexes

We have investigated the complexes formed by oligonucleotides with the general sequence d(T15,Gn), where n = 4-15. Two distinct classes of structures are formed, namely, the four-stranded tetraplex and frayed wires. Frayed wires differ from four-stranded tetraplexes in both strand association stoichiometry and the ability of dimethyl sulfate to methylate the N7 position of guanine. Thus, it appears that these two guanine-rich multistranded assemblies are stabilised by different guanine-guanine interactions. The number of contiguous guanine residues determines which of the complexes is favoured. Based on the stoichiometry of the associated species and the accessibility of the N7 position of guanine to methylation we have found that oligonucleotides with smaller number of contiguous guanines; n = 5-8, form primarily four-stranded tetraplex. Oligonucleotides with larger numbers of contiguous guanines adapt primarily the frayed wire structure. The stability of the complexes formed by this series of oligonucleotides is determined by the number and arrangement of the guanines within the sequences. We propose that the formation of the two types of complex proceed by a parallel reaction pathways that may share common intermediates.     

Borate complexes of amphotericin B: Polymeric species and aggregates in aqueous solutions

Amphotericin B, a polyene macrolide antibiotic, exists in aqueous solution as a poorly soluble, high-molecular-weight aggregate. A borate complex of this polyene was prepared that has greater solubility and is less aggregated. In aqueous solution this borate complex exists as a mixture of several molecular species differing in borate content, molecular weight, and molecular conformation. The solubility varied with pH and was minimal at neutrality. Throughout the pH range it was one to two orders of magnitude higher than that of the parent compound. The molecular size distribution, as determined by differential ultrafiltration, showed a progressive increase in the weight fraction of aggregates going from acid to alkaline solutions. The sizes of aggregates ranged from under 25 to over 100 molecules. The borate content of the complexes increased with increasing pH. No borate was complexed in acid solutions. This indicated that amphotericin B and borate ions can complex to form copolymer chains of varying length in which these species alternate, since both are bifunctional. The complexation equilibrium is favored by high pH. Absorption and CD spectra indicated that the polyene molecules can stack reversibly to form dimers. Dimerization constants calculated from the spectra were highest in neutral solution and declined with increasing acidity or alkalinity. In alkaline solutions the polymer chains are long and extended, with minimal stacking. In neutral solution the chains are shorter and extensively stacked. In acid solutions no borate complexes are formed, and the polyenes are stacked to an intermediate degree. The very different effects of pH and concentration on the degree of complexation with borate and on the degree of dimerization of the polyenes shows that these equilibria are independent of each other.     

Formation and characterization of soluble complexes of histone H1 with supercoiled DNA

We have analyzed the interaction of rat liver histone H1 with superhelical DNA. Depending on the ratio of H1 to DNA and the concentration of salt, two different types of complexes were found. Above a critical ratio of H1 to DNA, called the aggregation point, large aggregates are formed, which have a cable-like appearance in the electron microscope. Below the aggregation point, individual soluble complexes are formed, which are the subject of this study. With increasing ionic strength, the aggregation point is shifted towards lower ratios of H1 to DNA. In the soluble complexes, H1 appears to bind along superhelically intertwined DNA strands, forming a polymer. Partial digestion of the complexes with protease suggests protection of the N-terminal tail and the globular domain of H1. Similar soluble complexes were observed with various H1 fragments but not with the core histones. In the soluble complexes, similar regions of the H1 molecule are considered to be protected from cleavage by protease, as in chromatin. Therefore, these complexes appear to be a valuable model for the interaction of H1 in chromatin fibers.     

Sodium ion and solvent nuclear relaxation results in aqueous solutions of DNA

The field-dependent ²³Na nuclear relaxation in aqueous DNA solutions has been obtained for a range of temperatures, including the DNA melting region. At least two correlation times are needed to characterize the spectral density function for the ²³Na relaxation. For the slow process (with the largest correlation time), the temperature dependence of the coupling constant and the correlation time were determined, and important premelting effects were observed. Possible origins of the slow process are discussed. The last process is shown to be correlated with the properties of the hydration water of DNA as reflected by the ¹⁷O relaxation rates in these solutions. The influence of the polyelectrolyte and NaCl concentrations on the ²³Na relaxation rate is compared with previous results from solutions of linear flexible polyelectrolytes.     

The DNA melting transition in aqueous magnesium salt solutions.

The melting transition of the magnesium salt of DNA has been systematically examined in the presence of various types of anions. The addition of ClO4- to a concentration of 3.0 N results in the biphasic optical transition, with the first phase exhibiting rapid reversibility and independence of the DNA concentration. This subtransition, which is interpreted as an intramolecular condensation to a collapsed form of DNA, is followed by a DNA concentration-dependent aggregation reaction. The aggregation can be reversed by increasing the ClO4- concentration to 6.0 N while elevating the temperature to post-transition levels. Alternatively, both the collapse and the aggregation can be prevented by melting in the presence of trichloroacetate, the most strongly chaotropic solvent for DNA which has been reported (K. Hamaguchi and E. P. Geiduschek (1962), J. Am. Chem. Soc. 84, 1329). The forces responsible for mediating both the collapse and the aggregation are superficially similar to those involved in maintaining duplex stability. The collapsed form, in particular, possibly possesses features in common with the condensed structures which can be produced in aqueous solution of certain polymers, such as polyethylene glycol (Lerman, L.S. (1971), Proc. Natl. Acad. Sci. U.S.A. 68, 1886).     

Formation of free radicals and DNA breaks after pulsed laser irradiation of DNA complexes with intercalating dyes

Destruction of lambda phage DNA is studied under nanosecond pulse laser irradiation (lambda = 355 nm) of DNA-dye complexes in solution at 77K (dye--acridine orange, 8-methoxypsoralen, ethidium bromide). Free radicals induced by laser radiation are found to participate in DNA sugar-phosphate chain scission. It was observed that the quantity of DNA double-strand breaks correlated with that of the free radicals and that of oxygen influenced DNA laser destruction.     

On the mechanism of dielectric relaxation in aqueous DNA solutions.

The complex dielectric response of calf thymus DNA in aqueous saline solutions has been measured from 1 MHz to 1 GHz. The results are presented in terms of the relaxation of the incremental contributions to the permittivity and conductivity from the condensed counterions surrounding the DNA molecules. Measurements of the low-frequency conductivity of the samples also lends support to the condensed counterion interpretation.     

DNA conformation and thermostability of aqueous solutions of beta-alanine and gamma-aminobutyric acid

It has been shown that at low concentrations of rare amino acids (from 10(-3) M to 10(-1) M of the substance) stechiometric complexes amino acid -- DNA are formed, which bring about partial substitution of counterions screening phosphate groups and to a change of spatial structure of DNA water molecules. The DNA-solvent molecular interactions are changed, accompanied by an abrupt decrease of helix-coil enthalpy transition which leads to the unwinding of DNA double helix. In the region of amino acid high concentrations (greater than 1-1,5 M) a rise of thermostability and winding of DNA double helix is observed. It has been established that B----C-like conformational transition stimulated by the rise of DNA thermostability is a result of counterions dehydration and the increase of effective ionic strength of the solution which is due to the rise of amino acid-zwitterions content in it.