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Biomedical subjects

L I Shabarchina

Publications and source records attributed to L I Shabarchina.

At least 19 recordsLinked to original sources

Multilayer films containing immobilized nucleic acids. Their structure and possibilities in biosensor applications.

Langmuir-Blodgett (LB) and film technologies based on electrostatic attraction self-assembly (SA) are shown to be useful for immobilization of nucleic acids (DNA, polynucleotides) onto solid supports in sensor devices. The nucleic acids were immobilized in complexes with cationic surfactants (for LB) and polycations (for SA). Infrared spectral studies showed that DNA unfolds in multilayer LB films with octadecylamine and conserves its double helical structure in the LB films with dioctadecyldimethylammonium and in the SA films with polyallylamine, polyethylenimine and poly-L-lysine. Atomic groups and the types of interactions determining the complex formation of these films have been identified. The hydration of LB and SA films was studied to find out binding sites of water molecules and to evaluate the flexibility of nucleic acid compounds in the multilayer films. The possibilities of biosensor applications of these LB and SA films were monitored on binding of specific reagents for DNA by DNA-containing films and mononucleotides by a complementary single-stranded polynucleotide immobilized on a positively charged solid support.

Biosensing Techniques↗

[Irreversible changes in phage DNA after its protonation in solution and inside the virion detected by the transfection method].

The dependence of irreversible structural changes in phage lambda DNA on the degree of its protonation in a solution and inside the virion has been found by measuring the transfection activity of bacteriophage. The different effect of ionic strength on pH-dependence of the irreversible changes in the structure of DNA upon its protonation in a solution or in situ has been registered and explained. The insignificant shift of pH from neutral region value in 0.1 M NaCl has resulted in a damaging effect of H+ ions on compact DNA in situ as compared to the DNA in a solution. The effect of H+ ions on compact DNA in situ is mainly based on the formation of noncovalent intermolecular DNA-protein and DNA-DNA linkages.

Bacteriophage lambda↗

[Lactate dehydrogenase in an interpolyelectrolyte complex. Function and stability].

A new method for encapsulating enzymes by multilayer polyelectrolyte coating is proposed. The method consists in a stepwise adsorption of polyelectrolytes from solution onto protein aggregates formed by salting out the proteins in highly concentrated salt solutions. Polystyrene sulfonate and fluorescence-labeled polyalylamine were used for capsule formation. The size of lactate dehydrogenase aggregates covered by four layer pairs of electrolytes was 1-5 microns, as indicated by fluorescence microscopy. The catalytic characteristics and stability of pig muscle lactate dehydrogenase (EC 1.1.1.13) incapsulated in multilayer electrolyte complex obtained by this method were studied. It was found that the affinity of the substrate pyruvate for the enzyme in the polyelectrolyte complex (K(M)) did not essentially change as compared with the free enzyme. Incapsulated lactate dehydrogenase showed the following features that distinguish it from the free form: (1) the lifetime in diluted solutions increases from 30 min (without capsules) to 1-2 days (in capsules); (2) a higher stability to basic denaturation (up to pH 10); and (3) the absence of substrate inhibition of enzyme in the polyelectrolyte complex. The changes in the catalytic characteristics of incapsulated lactate dehydrogenase are discussed in terms of an increase in effective pK values of amino acid perturbed by polyelectrolyte coating of enzyme.

Animals↗

[Conformation and molecular and ionic transformations of polycytidylic acid immobilized in multilayer Langmuir and polyelectrolyte films].

Multilayer films of complexes of polycytidylic acid with dioctadecyldimetylammonium were obtained by the Langmuir-Blodgett method (LB films), and complexes of poly(C) with polycations (poly-L-lysisne, polyethyleneimine, polyallylamine) were obtained by the method of alternate adsorption (polyionic assembly) from solutions of oppositely charged polyelectrolytes on the solid carrier (SA films). It was shown that poly(C) exists in SA films in a single-stranded state irrespective of whether in the starting solution it occurred in the single-stranded nonprotonated or double-stranded protonated conformation. Conversely, in the LB film poly(C) preferred to be in a double protonated conformation. UV-spectra of water-insoluble LB and SA films at different pH values of surrounding water medium were investigated. Proton titration curves of poly(C) immobilized in LB films were obtained. The analysis of the shape of titration curves showed that the molecular-ionic transformation of poly(C) in LB films is accompanied by both the conformational transition of the polynucleotide and the molecular rearrangement in the whole film. Poly(C) was found to transform from the double- to single-stranded state and vice versa in the "deprotonation-protonation" cycle of LB film due to cooperative release/binding of hydrogen ions by cytosine bases. In contrast, poly(C) "protonation-deprotonation" in SA films occurred without conformational transitions of the polynucleotide. As opposed to poly(C) in solution a rather big hysteresis of forward and back titration curves was found for both types of multilayer films, indicating molecular rearrangements in films. The reason for the structural transformations of poly(C) upon fabrication of LB or SA films and the mechanism of molecular ionic transformations of poly(C) in films are discussed in terms of a simple model of ion exchange. An assumption about the nature of structural transformations of LB and SA films during their protonation-deprotonation is put forward.

Electrolytes↗

[Optical properties and molecular structure of nucleic acids and their components. X. Infrared spectra and molecular structure of denatured DNA with different water content and degree of protonation].

IR-spectra (4000-900 cm-minus-1) of heat denatured DNA (d-DNA) of calf thymus have been obtained in the range of relative humidity of 0-93% and of the guota of protoned bases alpha of 0-0,5. fron these spectra the diagram of d-DNA states is plotted as the relationships of intensities and the location of some bands with and relative humidity. At alpha approximately 0,2 a compact structure S2-1 with firm hydrogen bonds between protoned and unprotoned bases which is similar to those observed earlier while protoning homopolynucleotides poly-A and poly-C. At G1'-large values of alpha S2-1-conformation destroyed and a loose structure (alpha approximately 0,35) appears. the latter is characterized by a minimum H-binding of the base and increased hydration. Further increase of alpha brings about the formation of sufficiently compact G2-1-STRUCTURE WHICH IS CHARACTERIZED BY SOLID H-bonds between PO2-groups and protoned bases. A specific effect of the structures found on the state of absorbed water and the structure of hydrate envelope is revealed.

Animals↗

[Spectrophotometric study of the system DNA-lipid].

The effect of some physical and chemical factors on the system DNA-lipid and its components was studied by spectrophotometry. A change of optical density (D) of watersalt suspensions of lipid total fraction in the range of wavelengths from 200 to 350 nm with the change of ionic strength from 0 to 1 M NaC1 and temperature from 20 to 100 degrees C was investigated. Measurements were carried out under various temperature regemes. In all cases a bell-like irreversible dependence of D on temperature was observed. Its parameters were essentially dependent on the temperature heating of the samples and medium composition. In the presence of DNA the parameters of the curve were changed significantly and depended on the ratio DNA:lipid. An inhibiting effect of DNA on the oxidation of lipids and stabilization of liposomes as well as thermic destruction of DNA stabilized by lipids were observed under certain conditions. The data obtained allowed a conclusion concerning the formation of lipodesoxinucleic complex. It was shown that when a large number of lipid oxidation products was accumulated in the system a chemical modification of DNA with the change of its secondary structure went on.

Animals↗

[DNA-protein cross-links as a possible reason for genomic damage during its protonation].

The change in survival of bacteriophages with DNA of different GC-contents after their incubation in media of different acidities with subsequent neutralization was studied. It was shown that the higher the GC-content, the more sensitive is the phage to the action of H(+)-ions. Evidence is presented that the acidic inactivation of virions is not connected with the helix-coil transition of the intraphage DNA due to its protonation. The extractability of DNA from phages subjected to different concentrations of H(+)-ions with subsequent neutralization of the medium to pH 8 was determined. The changes in: transfection ability, UV-spectra, the quantity of the residual proteins, and the contents of glutamic and lysine amino acid residues in these proteins were investigated. The effect of glutamic acid on the parameters of DNA melting curves was followed for different pH values. Proceeding from the data obtained, we concluded that acidification of the medium from neutral tp pH congruent to 4 leads to formation of non-covalent DNA-protein cross-links due to interaction of the GC base pairs of DNA with glutamic and aspartic amino acid residues, whereas acidification of the medium to pH less than 4 with subsequent neutralization to pH 8 results in the formation of covalent DNA-protein cross-links of Schiff base type. The influence of non-covalent DNA-protein cross-links on the properties of DNA and their regulatory role in genome functioning are discussed.

Bacteriophage lambda↗

[The role of protons in a medium in gamma-radiation-induced damage of nucleic acids].

The study of the combined effect of gamma-radiation and acid medium (pH 7.0-2.0) on DNA and RNA showed that the radiation-induced injury to nucleic acids increased with increasing concentration of H+-ions in the medium up to pH values below which protons exerted a protective action. Irradiation of native DNA in acid medium, as compared to neutral one, increased not only the number of injured bases but also the average size of the induced local defect in the secondary structure. It was shown that the proton sensitization was determined both by the number of protonated bases and by the degree of ordering the polynucleotide chain.

Animals↗

[IR-spectroscopic study of the temperature anomaly of water sorbed onto DNA].

IR-spectra of liquid water and that sorbed on native and denaturated DNA at different relative humidity (RH) are obtained in the region of valent OH-(OD)-oscillations within the temperature range 0--100 degrees C. An S-shape decrease of absorption intensity was observed for water sorbed on DNA at 94% RH in two temperature regions (25--45 and 75--95 degrees C). Proceeding from the assumption that individual absorption bands are described by equal symmetrical functions, development of complex absorption contour of D2O in the region 3000--2000 cm-1 into constituents was performed. It enabled to determine band parameters of underphase vuf and synphase vsf valent and overtone of deformation oscillations of OD-groups in liquid and sorbed water and to reveal at higher temperatures the bands of "free" OD-groups (v = 2668 cm-1). Relative contribution of integral intensity of the latter at 96 degrees C in sorbed water is about 4 times lower than in the liquid one.

Chemical Phenomena↗

[Study of structural transformations of the sugar-phosphate chain and nitrogen bases of DNA hydrate by IR-spectroscopy].

The IR-spectra (900-1800 cm-1) of films of native and denatured DNA were studied within the temperature range 4-100 degrees C at different relative humidity (r.h.). Two temperature intervals of S-like change of spectral parameters of absorption bands of sugar-phosphate chain of double-helical DNA are found on temperature relationships at high humidity. These two intervals point to two types of cooperative transitions of sugar-phosphate pattern. The band parameters of nitrous bases are changed only under high temperature transition. In the region of structural transitions of different subsystems of DNA hydrate hyperchromism was wound for the bands of sugar-phosphate chain and hydrate cover unlike hyperchromism specific of most nitrous bases bands. From the data obtained it is proposed to use the basic characteristics of temperature relationship of spectral parameters of the absorption bands of sugar-phosphate pattern and hydrate envelope in the region of low-temperature transition for the "in vivo" analysis of the DNA structure in complex biological systems.

Chemical Phenomena↗

[Infrared spectroscopic study of DNA--lipid interactions. DNA compacting on disperse particles].

Within the range of relative humidity (r. h.) 0 to 92% there were obtained IR-spectra (4000--900 cm-1) of undeuterated and deuterated films of rat liver lipids, both in the free state and in the complex with native (nDNA) and heat-denaturated DNA (dDNA). Found peculiarities of IR-spectroscopic realization of the complex-formation are explained qualitatively. One of the peculiarities is hyperchromism of lipid bands and hypochromism of the bands of DNA bases. Lipids are shown to despiralize nDNA in the course of their interaction. A suggestion is advanced concerning the nature of basic contacts which form the complex DNA-lipid. It is found that nDNA unlike dDNA produces an inhibiting effect on lipids oxidation and their subsequent hydrolysis. The mechanism of the phenomenon is discussed. The ideas on DNA compactization resulting from its coiling on different protein and nonprotein particles are developed.

Animals↗

[Structure and function of DNA--membrane contact in cells].

A model of DNA-membrane contact based on the postulate about the determining role of membrane RNA (mbRNA) in providing its specifitity, stability and functional activity is proposed. Proceeding from the DNA-membrane contact structure, unity and interconditionality of reduplication and transcription processes are shown, and basic peculiarities of reduplication are discussed. It follows from the analysis of lysozyme-detergent treatment that two different DNA-membrane complex preparations can be obtained: one of these upon the spheroplast - "turned inside out spheroplast" transition, the other, containing specific DNA-membrane contact only - in the course of further detergent treatment. The parameters of the spheroplast - "turned inside out spheroplast" transition can be used to characterize the native DNA-membrane complex. It is concluded that RNA that stabilizes and makes the compact form of bacterial nucletide (f-DNA) is none other than the postulated mbRNA of the cellular DNA-membrane contact.

Bacteria↗

[The causes of hydration changes during DNA protonation].

Changes of DNA hydration provoked by protonation in the way of Na+- and H+-ions exchange, and in the way of HCl addition to Na+-DNA, were analysed by IR-spectroscopy. Water is shown not to contribute essentially to the formation and stabilization of conformations arising when DNA is protonated. The differences between hydratation of DNA protonated by different ways are in the main accounted for by alteration of the quantities of Na+ and Cl- ions forming the aqueous-salt envelope of polynucleotide.

Animals↗

[Mechanism of the radiation damage to the secondary structure of DNA].

The authors describe the mechanism of radiation-induced damage of a secondary DNA structure suggesting that a double-helical brim conformation adjacent to a local defect in the secondary structure, and the thermoactivated threshold process of a "confluence" of the defects are present in the irradiated DNA. It is supposed that the major type of the radiation damage to the spatial DNA structure is a triple helix repaired with the aid of the specific endonuclease.

DNA↗