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.     

Resistivity of Red Blood Cells Against High-Intensity, Short-Duration Electric Field Pulses Induced by Chelating Agents

The interaction of human red blood cells (RBCs) with diethylenetriamine-pentaacetic acid (DTPA) or its Gd-complex (Magnevist, a widely used clinical magnetic resonance contrast agent containing free DTPA ligands) led to the following, obviously interrelated phenomena. (i) Both compounds protected erythrocytes against electrohemolysis in isotonic solutions caused by a high-intensity DC electric field pulse. (ii) The inhibition of electrohemolysis was observed only when cells were electropulsed in low-conductivity solutions. (iii) The uptake of Gd-DTPA by electropulsed RBCs was relatively low. (iv) (Gd-) DTPA reduced markedly deformability of erythrocytes, as revealed by the electrodeformation experiments using high-frequency electric fields. Taken together, the results indicate that (Gd-) DTPA produce stiffer erythrocytes that are more resistant to electric field exposure. The observed effects of the chelating agents on the mechanical properties and the electropermeabilization of RBCs must have an origin in molecular changes of the bilayer or membrane-coupled cytoskeleton, which, in turn, appear to result from an alteration of the ionic equilibrium (e.g., Ca²⁺ sequestration) in the vicinity of the cell membrane. Received: 19 January 1999/Revised: 1 April 1999     

Sorption properties of polystyrene plates used in immunoenzyme analyses

The capacity of polystyrene carriers used in the enzyme immunoassay (EIA) for adsorbing 131I-labeled human serum albumin under different conditions has been studied, and the comparison of the plates manufactured by Dynatech AG (Switzerland) and by the Leningrad Works of Medical Polymers has been made. At the first stages of the reaction the antigen is separated from the carrier and the amount of the desorbed antigen depends on its initial dose and the dilution of the assayed sera. The irregular desorption of the antigen leads to misinterpretation of the results. Comparison of the polystyrene plates has shown that each plate is characterized by individual adsorption capacity, which impedes at present the standardization of EIA-based test systems.     

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.     

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.     

The Role of Mitochondrial DNA Individuality in the Pathogenesis of Parkinson’s Disease

Russian Journal of Genetics - The present article reviews the rapidly growing body of research on the role of mitochondrial DNA (mtDNA) in the realization of individual risk of Parkinson’s...     

Protein-filled polyelectrolyte microcapsules in the design of enzymic microdiagnostics

Encapsulation of enzymes (lactate dehydrogenase and urease) in polyelectrolyte shells was assessed with a view to designing enzymic microdiagnostics for low-molecular compounds in native biological fluids. Polyelectrolyte microcapsules were prepared with two polyanions [poly(styrenesulfonate) PSS and dextran sulfate DS] and two polycations [poly(allylamine) PAA and poly(diallyldimethylammonium) PDADMA]; calcium carbonate microspherulites with embedded enzymes served as “cores.” It was demonstrated that the main problem in making such a biosensor is to select a pair of oppositely charged polyelectrolytes that would be optimal for enzyme functioning. The best pairs were PAA/DS and PAA/PSS for lactate dehydrogenase, and PSS/PAA and PSS/PDADMA for urease. We designed and prepared enzyme-containing microcapsules differing in polyelectrolyte composition and number of layers, and investigated their properties.     

Hypotonic Activation of the Myo-Inositol Transporter SLC5A3 in HEK293 Cells Probed by Cell Volumetry,Confocal and Super-Resolution Microscopy

Swelling-activated pathways for myo-inositol, one of the most abundant organic osmolytes in mammalian cells, have not yet been identified. The present study explores the SLC5A3 protein as a possible transporter of myo-inositol in hyponically swollen HEK293 cells. To address this issue, we examined the relationship between the hypotonicity-induced changes in plasma membrane permeability to myo-inositol P _ino [m/s] and expression/localization of SLC5A3. P _ino values were determined by cell volumetry over a wide tonicity range (100–275 mOsm) in myo-inositol-substituted solutions. While being negligible under mild hypotonicity (200–275 mOsm), P _ino grew rapidly at osmolalities below 200 mOsm to reach a maximum of ∼3 nm/s at 100–125 mOsm, as indicated by fast cell swelling due to myo-inositol influx. The increase in P _ino resulted most likely from the hypotonicity-mediated incorporation of cytosolic SLC5A3 into the plasma membrane, as revealed by confocal fluorescence microscopy of cells expressing EGFP-tagged SLC5A3 and super-resolution imaging of immunostained SLC5A3 by direct stochastic optical reconstruction microscopy (dSTORM). dSTORM in hypotonic cells revealed a surface density of membrane-associated SLC5A3 proteins of 200–2000 localizations/μm². Assuming SLC5A3 to be the major path for myo-inositol, a turnover rate of 80–800 myo-inositol molecules per second for a single transporter protein was estimated from combined volumetric and dSTORM data. Hypotonic stress also caused a significant upregulation of SLC5A3 gene expression as detected by semiquantitative RT-PCR and Western blot analysis. In summary, our data provide first evidence for swelling-mediated activation of SLC5A3 thus suggesting a functional role of this transporter in hypotonic volume regulation of mammalian cells.