[A device to control the mass concentration of protein-containing aerosoles in real-time mode].

The article is dedicated to the device to measure the mass concentration of protein-containing aerosols, which is now being developed at State Scientific and Research Institute of Biological Instrument Making. The operation of the device is based on the registration of spectral-luminescent and dispersing properties of individual particles of biological nature. Experiments carried out on aerosol static-dynamic stand have demonstrated that the device measures the mass concentration of protein-vitamin concentrate aerosol in real-time mode with 50% accuracy.

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.

Fluorescence quenching and spin label electron spin resonance studies of stacking self-association in aqueous solutions of 2-aminopurine riboside and its 5'-mono- and -diphosphate.

The autoassociation of 2-aminopurine riboside (rn2Pur) and its 5'-mono- (P-rn2Pur) and 5'-diphosphate (PP-rn2Pur) in neutral aqueous solutions was investigated using fluorescence quenching and ESR spin-label methods within the range 276-358 K. Respective equilibrium constants and thermodynamic functions were derived therefrom assuming two models of infinite autoassociation: (i) an isodesmic one (K2 = K3 = ... Kp), and (ii) one in which K2 no equal to K2 = K4 ... Kp. Comparative analysis of these data and that of the parent 2-aminopurine, obtained previously, allowed us to formulate the following conclusions: (1) the mechanism of autoassociation of rn2Pur varies with temperature in such a way that a T = 318 K the isodesmic model is fulfilled (K2 = Kp); at high temperatures Kp/K2 greater than 1, i.e. the process is cooperative, while at lower temperatures it becomes anticooperative (Kp/K2 greater less than 1); (2) at 298 K the tendency to autoassociation decreases in the order; rn2Pur greater than P-rn2Pur greater than PP-rn2Pur; (3) rn2Pur forms highly packed complexes with the bases stacked and the ribofuranose residues interacting via hydrogen bonds or water bridges; (4) autoassociation of P-rn2Pur and PP-rn2Pur is mainly governed by stacking of the bases, while the ribose phosphate residues attain a trans configuration corresponding to the lowest electrostatic repulsion between charged phosphate groups; even at high ionic strength (I = 0.8), a positive electrostatic contribution to the free enthalpy of autoassociation is observed; (5) the two methods employed gave similar results for P-rn2Pur, but somewhat different ones for rn2Pur because the presence of the spin label (nitroxide stable radical) at the 2'(3')-OH group of the ribose residues prevents its interaction via hydrogen bonding with an unlabeled one of an adjacent nucleoside.

Structural and thermodynamic features of complexes formed by DNA and synthetic polynucleotides with dodecylamine and dodecyltrimethylammonium bromide.

Complex formation of native and denatured DNA, single-stranded polyribonucleotides poly(A) and poly(U), as well as double-stranded poly(A).poly(U) with dodecylamine (DDA) and dodecyltrimethylammonium bromide (DTAB) has been studied by UV-, CD-, IR-spectroscopy and fluorescence analysis of hydrophobic probe pyrene. DDA and DTAB were shown to bind cooperatively with DNA and polyribonucleotides, resulting in the formation of complexes containing hydrophobic micelle-like clusters. Critical aggregation concentration (CAC) of DDA and DTAB shifts sharply to lower values (30-50 times) in the presence of DNA and polynucleotides as compared to critical micelle concentration (CMC) of free DDA and DTAB in solution. The analysis of binding isotherms within the frame of the model of cooperative binding of low-molecular ligands to linear polymers allowed us to determine the thermodynamic parameters of complex formation and estimate the contribution of electrostatic interaction of positively charged heads of amphiphiles with negatively charged phosphate groups of DNA and polyribonucleotides, and hydrophobic interaction of aliphatic chains to complex stability. Electrostatic interaction was shown to make the main contribution to the stability of DNA complexes with DDA, while preferential contribution of hydrophobic interactions is characteristic of DTAB complexes with DNA. The opposite effect of DDA and DTAB on the thermal stability of DNA double helix was demonstrated from UV-melting of DNA-while DTAB stabilizes the DNA helix, DDA, to the contrary, destabilizes it. The destabilizing effect of DDA seems to originate from the displacement of intramolecular hydrogen bonds in complementary Watson-Crick A.T and G.C base pairs with intermolecular H-bonds between unsubstituted DDA amino groups and proton-accepting sites of nucleic bases.

[Kinetics of formation of 8-oxy-2'-deoxyguanosine-5'-monophosphate under the effect of heat: determination of rate constants and activation energy]. / Kinetika obrazovaniia 8-oksi-2'-dezoksiguanozin-5'-monofosfata pod vliianiem tepla: opredelinie konstant skorostei i énergii aktivatsii.

Rate constants of 8-oxy-dGMP (8-hydroxy-dGMP) formation upon incubating dGMP in H2O solutions at different temperatures were determined with differential UV-spectroscopy. Extrapolation of rate constant values obtained at elevated temperatures to 37 degrees C gives k = 5.8 x 10(-10) s-1.M-1. The activation energy for the process was estimated to be 24 kcal/mole. In D2O solutions essential lowering of the activation energy (13 kcal/mole) and rising of rate constant (k = 3.7 x 10(-9) s-1.M-1 at 37 degrees C) were observed. The strong influence of D2O on the process points to the possible participation of singlet oxygen in a heat-induced formation of 8-oxy-dGMP. The obtained values of rate constants and activation energy induced by heat show that of all types of DNA damages currently known such as single strand scission, depurination, cytosine deamination and oxidation of guanyl residues to the 8-oxo-derivatives- the last process seems to be the strongest damage of DNA resulting in such biological consequences as mutagenesis, carcinogenesis and aging.

[Study of intermolecular interactions and self-organization of adenylic nucleotides by the spin label method]. / Issledovanie mezhmolekuliarnykh vzaimodeistvii i samoorganizatsii adenilovykh nukleotidov metodom spinovykh metok.

A spin labeling method for obtaining thermodynamic parameters of nucleotide association is proposed. The method is based on the dependence of ESR parameters of the spin-labelled derivative on concentration of the nonlabelled compound due to formation of associates involving both spin-labelled and unmodified molecules. It is found that at pH 7.5 the constant of adenylic nucleotide association practically does not depend on the number of phosphate groups and is equal to 9.7 +/- 0.3 M-1 for AMP, ADP and ATP in 0.1 M NaCl at 28 degrees C. In acidic medium the value of the association constant increases by a factor two. Base stacking is shown to make the main contribution to stability of the associates of adenylic nucleotides at neutral pH, whereas upon base protonation the key role is played, apparently, by base-phosphate interaction. It is thought, that an increase in the solvent entropy is essential for stabilisation of the associates, this factor being more important in the case of nucleotide association as compared to the association of nucleosides. A possible role of nucleotide association in the processes of intracellular regulation is discussed.

[Segmental flexibility of single-, double-, and triple-stranded polyribonucleotides from the data of spin label method. Formation of the triple-stranded poly(A.A.U) polynucleotide helix]. / Segmental'naia gibkoct' odno-, dvukh- i trekhspiral'nykh poliribonukleotidov po dannym metoda spin-metki. Obrazovanie trekhnitchatoi polinukleotidnoi spirali poli(A.A.U).

Segmental mobility dynamic peculiarities of poly(U), poly(A) and poly(C) synthetic polymers and their complexes were investigated by spin-label method. Imidazolide spin-label was introduced into 2'-oxi-groups of polymer ribose in correlation: one spin-label on 18-20 bases. Formation of complexes was observed by ESR spectra at two pH: 4.2 and 7.2. Segmental mobility of only single strand spin-labelled polymer segment and in the complex was evaluated by measuring rotational correlation time (tau) determined by dependence of distances between outer wide extrema in ESR spectra from solvent viscosity at different temperatures. It turned out that correlation time tau of single strand structures in a high degree depend on pH and temperature. For three strand structures abrupt increase of tau because of appearance of rigidity was observed. It is possible to evaluate part of triple complexes poly(U.A.A) and poly(U.U.A) existing in dynamic equilibrium depending on pH and temperature by the form of outer wide extrema. Adding of dye to complex of poly(U).poly(A) causes an increase of rigidity of the supermolecular structure. Quantitative characteristics of formed complexes were obtained by simulation of ESR spectra on computer.

[DNA-protein cross-links as a possible reason for genomic damage during its protonation]. / DNK-belkovye sshivki kak vozmozhnaia prichina povrezhdenii genoma pri ego protonirovanii.

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.

[Complex formation between polyuridylic acid and adenosine using spin-labeling]. / Issledovanie kompleksoobrazovaniia mezhdu poliuridilovoi kislotoi i adenozinom metodom spinovykh metok.

2 poly(u) . 1 adenosine complex formation was studied by spin-technique in the temperature range from 7 to 25 degrees C. It is shown that adenosine binding induces the conformational transition of poly(u) structure from flexible coil into rigid helix. The contribution of hydrogen bonding and stacking of adenosine to the total change of enthalpy and entropy upon complex formation was estimated.

[Spin-labeled nucleosides, nucleoside-5'-mono-, di-, and triphosphates]. / Spin-mechenye nukleozidy, nukleozid-5'-mono-, di- i trifosfaty

A general method of synthesis of spin-labeled nucleosides, nucleoside-5'-mono-, di- and triphosphates is developed. It is based on the acyllation of ribose OH-group with N-(2,2,5,5-tetramethyl-3-carbonylpyrollin-1-oxyl)-imidasole. Their ESR and UV-spectra are studied. While studying the products of interaction between nucleoside mono-, di- and triphosphates and N-(2,2,5,5-tetramethyl-3-carbonylpyrrolin-1-oxyl)-imidasole along with the formation of monoradicals due to the acyllation of 2'(3')-OH-groups of ribose in the case of AMP, GMP and GDP acyllation by the phosphate group with the formation of biradicals has been also observed.

[Use of spin labels for studying the association of adenosine, adenosine-5'-mono-, di- and triphosphate in aqueous solutions]. / Primenenie spinovykh metok dlia issledovaniia assotsiatsii adenozina, adenozin-5'-mono-, di- i trifosfata v vodnykh rastvorakh.

Possibility of spin label studies as a method of investigation of different molecular associations is considered with adenosine, adenosine-5'-mono, di- and triphosphate as examples. The proposed method is based on dependence of the parameters of spin-labelled derivatives ESR-spectra on concentration of corresponding unlabelled compounds. Being highly sensitive the method takes the advantage of obtaining not only thermodynamic characteristics of association but dynamic parameters of molecular mobility of the associates as well. It is shown by this method that base-phosphate interaction participates in nucleotide association, together with the stacking interaction.

[Formation of complexes between DNA and cationic amphiphile molecules by the fluorescent probe method]. / Izuchenie kompleksoobrazovaniia DNK s kationnymi amfifil'nymi molekulami metodom fluorestsentnogo zonda.

The formation of complexes of DNA with dodecylamine, dodecyltrimethylammonium, tetradecyltrimethylammonium, and hexadecyltrimethylammonium was studied using a fluorescent probe pyrene. The dependences of the spectral parameters of the hydrophobic pyrene probe on the concentration of the cationic amphiphile in the presence and absence of DNA were obtained and analyzed. It is shown that, in the absence of DNA, these dependences exhibit only one S-shaped region, which corresponds to the micelle formation of the amphiphile, whereas in the presence of DNA there are two S-shaped regions, which indicates the cooperative formation of two types of DNA-cationic amphiphile complexes. For each of the four cationic amphiphiles, the critical concentrations for the micelle formation in the absence of DNA (C0) and the concentrations at which the first (Cd1) and the second complex with DNA are formed were determined. It was found that the Cd1 value is 15-40 times lower than C0. The Cd1 value does not depend on DNA concentration and is determined only by the length of the hydrocarbon chain and the structure of the amphiphile ionic fragment. The Cd1 value increases as the length of the aliphatic chain decreases and upon replacement of mobile hydrogen atoms in the ammonium fragment by methyl groups. It was shown that hydrophobic clusters of amphiphile arising upon complex formation with DNA play the role of cross-links promoting DNA aggregation, or DNA compactization in the case of dilute solution of high-molecular weight DNA. The structures of the first and second DNA-cationic amphiphile complexes are proposed, and the mechanism and nature of interactions that determine their formation are discussed.

[The dual stabilizing and destabilizing effect of decylamine on DNA structure and stability]. / Dvoistvennoe, stabiliziruiushchee i destabiliziruiushchee deistviie detsilamina na strukturu i stabil'nost' DNK.

The influence of decylamine on DNA structure and stability has been studied from the changes in UV-spectra and UV-melting curves. It was found that at low concentrations, the aliphatic amine stabilizes the double helix while at high concentrations, on the contrary, the amine disturbs it. The interaction of aliphatic amine with DNA results in a sharp increase in the heterogeneity of thermostability of different sites of the double helix. The stabilizing effect of aliphatic amines is connected with a decrease in repealing between negatively charged sugar-phosphate chains of the double helix, while the destabilizing effect is the result of displacement of intramolecular base-base hydrogen bonds by intermolecular base-amine H-bonds. The biological significance of the dual, stabilizing and destabilizing effects of aliphatic amines on DNA helix is shown.

[Remote monitoring control techniques for biological air disperse systems]. / Distantsionnye metody kontrolia aérodispersnykh sistem biologicheskogo proiskhozhdeniia.

Described in the paper are the results of designing a combination-luminescence lidar for the automated express determination of the concentration and dynamic distribution of biological pollutants in the atmosphere. It was established as possible to detect simultaneously the signals of aerosols' luminescence and those of combined atmospheric nitrogen dispersion to ensure an effective monitoring of biological aerosols along the probing route.

Complex formation between spin-labeled polyuridylic acid and pyrimidine nucleosides.

Complex formation between poly (U) and pyrimidine nucleosides, uridine and cytidine, was observed using spin labeling technique. The binding of these nucleosides with poly (U) takes place within a narrow range of their concentration and is characterized by a relatively strong cooperativity. It is shown, that both hydrogen bonding and stacking interaction contribute to the complex stability. Some thermodynamic parameters of the process were obtained from the binding isotherms. At 21 degrees C the equilibrium constants for nucleation were found to be 0.23 M-1 and 0.42 M-1, and those for chain growth 2.63 M-1 and 2.19 M-1 for uridine and cytidine respectively. Complex formation of poly (U) with adenosine was also studied by spin labeling method.

Spin-labeled polyribonucleotides.

Poly (U), poly (C) and poly (A) were spin labeled with N-(2,2,5,5-tetramethyl-3-carbonylpyrroline-1-oxyl)-imidazole. This spin label interacts selectively with 2' OH ribose groups of polynucleotides and does not modify the nucleic acid bases. The extent of spin labeling is not dependent upon the nature of the base and is entirely determined by rigidity of the secondary structure of the polynucleotide. The extent of modification for poly (U), poly (C) and poly (A) was 4.2, 1.7 and 1.5 per cent, respectively, the secondary structure of the polynucleotides being practically unchanged. Some physico-chemical properties of the spin-labeled polynucleotides were investigated by ESR spectroscopy. Rotational correlation times of the spin label and activation energy of its motion were calculated.

Laser beam propagation through the turbulent atmosphere with precipitation.

Equations of the Markov approximation method for the problem of laser beam propagation in the turbulent atmosphere with precipitation have been obtained by replacing precipitation particles with spatially disordered finite black screens and by replacing turbulent inhomogeneities with analogous phase screens. Solution of the equation for the mutual coherence function is represented as a convolution of the three functions using the Wigner function. The results of simultaneous measurements of laser irradiance fluctuation along the 200-m, 2.5-km, and 5-km paths during rain are presented. A qualitative model of irradiance fluctuations at multiple scattering owing to shading of a receiver by an adjacent particle layer is suggested. A saturation phenomenon of the laser radiation scintillation index in the turbulent atmosphere with precipitation is suggested and observed.

Probing of mRNA binding sites involved in interactions with rat liver ribosomes using poly(U) spin labeled at the ribose moiety.

The interaction of rat liver ribosomes with poly(U), spin labeled (SL) at the 2'OH groups of ribose residues by N-(2,2,5,5-tetramethyl-3-carbonylpyrroline-1-oxyl)-imidazole, has been studied by electron spin resonance (ESR) spectroscopy. The ESR spectra demonstrate that SL-poly(U) with a modification of 1 spin label per 20 ribose residues binds to 80S ribosomes as well as to 40S subunits in a 1:1 stoichiometry at 12 mM MgCl2. The same result is found with highly modified poly(U) bearing 1 SL per 4 ribose residues. Addition of excessive amounts of unmodified poly(U) displaces bound SL-poly(U) from the ribosome which points to a competition for the same binding site at the ribosome. The biological activity of SL-poly(U) was tested with regard to trigger 80S ribosomes for binding of Phe-tRNAPhe and for poly(Phe) synthesis. SL-poly(U) bearing 1 SL group per 20 ribose residues directs the binding of only 50% of the amount of Phe-tRNAPhe bound to ribosomes in the presence of unmodified poly(U). When SL-poly(U) bearing 1 SL group per 4 ribose residues is used, this value drops to 25%. Poly(Phe) synthesis is even more impaired: In the presence of poly(U) bearing 1 SL-group per 20 ribose residues only about 30% of the amount of poly(Phe) coded by unmodified poly(U) are synthesized and in the presence of SL-poly(U) bearing 1 SL group per 4 ribose residues poly(Phe) synthesis is completely abolished. The results suggest that modification of the ribose moiety has only a relatively small influence on the binding of mRNA to ribosomes but causes substantial impairment of the mRNA function, whereas, as shown earlier (Ebert et al., Acta Biol. Med. Germ. 41, 431, 1982), modification of the base moiety of poly(U) (in a proportion of 1 SL per 30 bases) does not influence coding efficiency for poly(Phe) synthesis.

[Interaction of a membrane preparation of Na+, K+-ATPase with a spin-labeled analog of ATP]. / Vzaimodeistvie membrannogo preparata Na+, K+-ATPazy so spin-mechenym analogom ATP

Interaction of membrane Na+, K+-ATPase preparation from brain gray matter with spin-labelled ATP analogue, in which free iminoxyl radical is joined as a result of 2'(3')-OH ribose groups acylation, is studied. The rotatory mobility of spin-labelled ATP analogue in Na+,K+-ATPase preparation is found to change in non-linear manner during temperature variation (the break-point on the curve being at 20-23degrees C). It correlates with temperature dependence of Na+,K+-ATPase and temperature dependence of lipid viscosity in the membranes, determined by means of hydrophobic spin probes. Substitution of Mg2+ ions with paramagnetic Mn2+ ions resulted in an intense magnetic dipole-dipole interaction between a spin label and Mn2+ ion, which indicated the formation of triple complex enzyme--spin-labelled ATP--Mn2+.