Том 61, № 6 (2016)

Proteins of the Dps family perform a dual function in bacterial cells. As ferritins, they protect cells from destructive effects of Fe²⁺ ions, while interacting with DNA they condense the genome in the absence of nutrients. The ability of Dps to self-aggregate is of a great importance. The way of genome remodelling from the condensed state to the active one is not yet known. Here, the effects of two sugar ligands on Dps interaction with DNA have been studied in vitro. For the first time it was demonstrated that D-glucuronate and D-galacturonate, but not D-glucose, can decompose the dodecameric structure of the protein and D-glucuronate stimulated the formation of binary complexes with the linear DNA fragments. As a result of flexible molecular docking, it was found that the molecules of all three sugars potentially can form clusters inside the protein cavity of Dps, but D-glucuronate and D-galacturonate were also bound in the region of intersubunit contacts of oligomer. The consequent destabilization of the intersubunit bonding network can, thus, be the main factor provoking the protein decay to the smaller oligomeric forms. Such a structural rearrangement, leading to a reduction in aggregation, may play a key role in genome decondensation during cell transition to the phase of rapid growth.

The possibility of increasing the effectiveness of antitumor drugs such as doxorubicin by preparing its complex with ultrafine magnetic iron oxide nanoparticles is considered. A method for binding doxorubicin molecules to magnetic nanoparticles via citric acid is proposed. The main magnetic properties of the obtained conjugates were studied by proton relaxometry and Mössbauer spectroscopy, while their cytotoxic activity was evaluated via spectrophotometric MTT assay in HeLa cells. It was shown that the conjugates of magnetite nanoparticles with doxorubicin are characterized by a high level of contrast in magnetic resonance imaging. The magnetic properties of doxorubicin-free and bound magnetite nanoparticles are mainly determined by the average size of nanoobjects and the phase composition and slightly depend on the composition of the stabilizing shell. The cytotoxic effect of the synthesized conjugates of magnetite nanoparticles with doxorubicin is higher than that of unbound doxorubicin. This makes it possible to increase the antitumor effect of doxorubicin and control the dynamics of its delivery in the form of a conjugate into the disease focus due to the magnetic contrast properties of nanoparticles.

The spatial organization of outer-membrane porins is studied by optical spectroscopy and molecular modeling. It was found that the OmpF and OmpC porins from Yеrsiniа ruckeri are β-structured membrane proteins typical of the pore-forming proteins of other Gram-negative bacteria. The spatial structures of monomers and trimers of the OmpC and OmpF porins from Y. ruckeri are simulated using methods of structural bioinformatics. It was found that the structural stability of the more thermostable OmpF trimer is sustained by a greater number of hydrogen bonds and hydrophobic interactions. The main differences of the spatial structures of the test porins are observed in the structure of their outer loops. There are three tryptophan residues in the molecules of the OmpC and OmpF porins of Y. ruckeri. It is demonstrated by moleculardynamics methods that after thermal denaturation the solvent accessibility of the Trp212 residue in OmpF porin increased by two times, while the solvent accessibility of a Trp184 residue in OmpC porin was not increased. It is hypothesized that the red-shifted tryptophan fluorescence spectrum of OmpF porin during thermal denaturation is due to the behavior of the Trp212 residue.

We have developed a technique for the immobilization of inulinase on chitosans with different molecular weights. The acid-soluble mid-molecular-weight (200 kDa) and high-molecular-weight (350 kDa) chitosans are shown to be promising matrices for inulinase adsorption. We assumed that the formation of an inulinase-chitosan matrix complex occurs mainly due to hydrophobic interactions; electrostatic interactions also play an essential role. The enzyme complex with high-molecular-weight chitosan is more stable against the action of UV light and temperature. This allows us to recommend chitosan as a catalyst in industry for the production of fructose from inulin-containing vegetable raw mateials.

We have studied the expression of cyclins (Cicl A and Cicl B1) and cyclin-dependent protein kinases (Cdk1, Cdk2, Cdk4, and Cdk5) in the brain of the long-tailed ground squirrel (Spermophilus undulatus) during different phases of their yearly cycle of life activities. We found that the expression of protein kinases in the frontal neocortex, hippocampus, and caudal brainstem differed by from three to five times, which indicates the regional specificity of the activity of cell-cycle proteins in the brain of a hibernating animal. During the end of winter hibernation, a significant increase in the expression of Cdk1, Cdk2, and Cdk4 were found in the hippocampus, which is due to the presence of progenitor neural cells in the subgranular region of the dentate gyrus. These cells are able to produce new neurons during all of ontogenesis. Our results show that during winter hibernation and awakening, region-specific changes in the expression of cell cycle proteins occur in the brain of a long-tail ground squirrel, which provides the appropriate activity of the cell cycle during the new functional state of a hibernating animal.

The effects of the drug flocalin, which possesses cardioprotective properties, on the respiration rates of rat-heart and liver mitochondria in different functional states, the efficiency of oxidative phosphorylation, as well as the transport of potassium ions in these organelles, were studied. It was found that flocalin at concentrations of 7–30 μm stimulated respiration of rat-heart and liver mitochondria in V₂ and V₄ states in the presence of succinic add as a respiration substrate in a potassium-containing medium. In the absence of potassium ions in the incubation medium, flocalin had no effect on mitochondrial respiration in these states. Studying the functioning of the potassium transport system revealed that flocalin at these concentrations dose-dependently activated the ATP-dependent transport of potassium ions in rat-heart and liver mitochondria. The data we obtained indicate that the cardioprotective effect of flocalin can be associated with activation of the ATP-dependent potassium channel of the inner mitochondrial membrane.

This paper presents the data and describes the Ca²⁺-dependent effect of the products of ω-oxidation of palmitic acid, as well as ω-hydroxypalmitic and α, ω-hexadecandioic acids, on rat erythrocytes. It is shown that in the presence of Ca²⁺ these acids induce aggregation of erythrocytes, which is accompanied by a reduction in the number of single cells in suspension. As well, a release of K⁺ from the cells occurs, which indicates the permeabilization of the plasma membrane. However, ω-hydroxypalmitic and α, ω-hexadecandioic acids are inferior to palmitic acid in their ability to induce Ca²⁺-dependent erythrocyte permeabilization. Bovine serum albumin and blood serum inhibit the effects of palmitic acid. At the same time, the influence of these agents on the effects of ω-hydroxypalmitic and α, ω-hexadecandioic acids appears to be much weaker. It is shown that ω-hydroxypalmitic and α, ω-hexadecandioic acids in the presence of Ca²⁺ induce an increase in the hydrodynamic diameter of single-walled lecithin liposomes, which indicates their fusion and (or) aggregation. The mechanisms of ω-hydroxypalmitic acid/Ca²⁺- and α, ω-hexadecandioic acid/Ca²⁺-induced effects on rat erythrocytes are discussed.

A model of the process of T-lymphocyte extravasation into a lymph node via the high endothelial venules in the course of the immune response has been developed. The histological structure and the morphometric parameters of the lymph node and its venules, as well as the presence of adhesion molecules on the endothelial cells and the speed of T-lymphocyte movement, were taken into account in the model and compared to the basic postulates of the clonal selection theory of immune surveillance. The inability of the venules of the lymph node to provide the passage of a sufficient number of T lymphocytes has been demonstrated; thus, the concept of immune surveillance formulated within the existing immunological theory has been proven inadequate. This finding points to the need for revision of the widely accepted concepts of the emergence of T-lymphocyte specificity and the very foundations of the clonal selection theory.

The temperature dependences of the charge-recombination rate in the ion-radical pair P870⁺Q_A^- in photosynthetic reaction centers of Rhodobacter sphaeroides were investigated. Recombination kinetics were measured in the individual absorption bands of the donor (600 nm) and an electron acceptor (335 and 420–450 nm) for the reaction center in the water–glycerol and trehalose environment after freezing preparations to–180°С in the dark and on the actinic light and after their subsequent heating. In similar conditions the fluorescence lifetime of tryptophanyls in reaction centers (λ_reg = 325 and 345 nm), which is an internal indicator of the dynamic state of the protein matrix, was measured. A correlation between the temperature dependences of functional and dynamic parameters of reaction centers in different solvents was shown. The differences in the average fluorescence lifetime of tryptophanyls in reaction centers of preparations frozen in the dark or on the actinic light were found. These results are explained due to transitions of reaction centers between different conformational states and processes of proton relaxation in the structure of the hydrogen bonds in the environment of reaction-center cofactors.

The Smoluchowski equation, which describes pore diffusion in the radius space, with a source term, is used in modeling the process of the formation of a hydrophilic pore in a lipid bilayer during phase transition. The introduction of a hydrophobic-pore source term into the equation reflects the emergence of additional defects in a bilayer caused by the decrease in the molecule area under the transition from the liquid crystalline to the gel phase. The distribution of the time probability density calculated within the model that is required for the formation of a hydrophilic pore is in good agreement with the previously published experimental data.

In vitro exposure of human erythrocytes to H₂O₂ at concentrations of 30–1000 μM resulted in a dose-dependent increase of the intracellular levels of Zn²⁺ and inhibition of the cytosolic esterase activity, which is a major marker of erythrocyte viability. The observed effect depended on the concentration of H₂O₂ and the duration of exposure of the cells to this compound. An inverse relationship between the changes in the intracellular level of labile zinc ions and esterase activity in the cells exposed to hydrogen peroxide was detected; this was indicative of the role of Zn²⁺ in the programmed death of red blood cells. The combined action of hydrogen peroxide and N',N'-tetrakis-(2-pyridyl-methyl)-ethylenediamine, an intracellular zinc ion chelator, has been found to eliminate the cytotoxic effect of H₂O₂, whereas the addition of Zn²⁺ to the erythrocyte incubation medium enhanced the effects of hydrogen peroxide. The reduction of the concentration of non-protein thiol groups due to a decrease of the level of reduced glutathione was shown to contribute to the release of Zn²⁺ from the intracellular binding sites during oxidative stress induced by H₂O₂ in human erythrocytes.

Mechanisms of tumor-cell responses to 2-isopropyl-5-methyl-1,4-benzoquinone (thymoquinone) and 1,4-benzoquinone were studied using fluorescence and the inhibition assay. It was shown that quinones enhanced the intracellular production of reactive oxygen species, reduced the mitochondrial membrane potential, and induced tumor-cell death through different pathways. It was found that thymoquinone, which induced lower production of reactive oxygen species than 1,4-benzoquinone, was more toxic to tumor cells. It was established that reactive oxygen species produced due to exposure to thymoquinone are involved in redox signaling processes that lead to the formation of mitochondrial permeability transition pores and activation of programmed cell death. These results suggest that the functioning of the established redox signaling mechanism is enabled by the colocalization of mitochondrial oxidoreductases involved in the production of reactive oxygen species and of protein targets of reactive oxygen species involved in the activation of apoptosis.

Low-intensity laser irradiation exerts effects on various biological objects and is currently exploited in various branches of medicine. From a practical point of view, irradiation in the near-infrared range seems most attractive since it has the highest penetration characteristics for human tissues. In the present work, we studied the effects of 835 nm infrared low-intensity laser irradiation on two melanoma cell lines, Mel IL and MeWo, in culture. The data that we obtained indicate that low-intensity infrared irradiation impacts the growth rate of Mel IL and MeWo melanoma cell lines and that the response dynamics of the two cell lines are similar, in spite of certain quantitative differences. Stimulation of cell growth occurs within a relatively narrow range of low doses (about 0.17 J/cm²). With significantly higher doses, deceleration of growth occurs instead. Рге-treatment of cells with low-dose radiation, however, protects them from the negative influence of higher doses. The protective action of low doses develops gradually, within about 10–30 min, and persists for at least 3 h. Рге-irradiation of cells also widens the range of stimulatory action with low-dose radiation. A theoretical model that explains the dynamics of cellular responses to various irradiation doses has been proposed.

The effects of continuous and amplitude-shift-keying low-intensity electromagnetic field (EMF) of the 10-m range with on–off cycling on fertility, offspring quality, and linear body size was studied in the planktonic crustacean Daphnia magna. Single exposure of 1-day-old crustaceans was shown to affect their reproductive characteristics, causing fetal abnormalities in the offspring and the linear dimensions of the body.

The procedure of differential tonometry by the Schiøtz tonometer is modeled using a two-component model of the eyeball in which the cornea is represented by a momentless deformable and linearly elastic surface and the sclera region by an elastic element that responds to changes in intraocular pressure via changes in volume. Numerical calculations showed that the dependence of intraocular pressure on the weight of a cornea- deforming rod (a plunger) is almost linear. This makes it possible to take the slope of its plot (pressure difference coefficient) as a characteristic of the eye. The coefficient is studied as a function of the elastic characteristics of the eye and the pressure in the unloaded eye. An analysis based on the dimensional theory and confirmed by calculations made it possible to conclude that in the first approximation the pressure-difference coefficient depends on the elastic constants through their ratios to the intraocular pressure. A comparison was made with the standard method of processing the data of differential tonometry.

The levels of large deletions in the mitochondrial DNA of workers at the Mayak Production Association (Mayak PA) who were exposed to external and combined occupational (external γ- and internal α-rays) radiation during the course of their duties were investigated. Peripheral blood-derived DNA samples were provided by the Radiobiological Human Tissue Repository of the Southern Urals Biophysics Institute (Russia). The samples were analyzed using long-extension PCR. The number of large-scale deletions in the mitochondrial DNA of workers who, in addition to external γ-radiation, were exposed to extra doses of irradiation due to incorporated ²³⁹Pu with a Pu body burden of 0.77–4.32 kBq, was 2.5-times lower compared to that of individuals who received only external γ-radiation. No significant gender-associated effects on the number of mitochondrial DNA deletions were detected among age-matched individuals.

We studied the effect of taurine on renal function and the prooxidant–antioxidant balance in the kidneys and blood of rats under experimental rhabdomyolysis. It was found that taurine at a dose of 100 mg/kg of animal body weight improves renal function and normalizes prooxidant–antioxidant balance indicators, reducing the intensity of lipid and protein peroxidation and increasing catalase and glutathione peroxidase activities.

A mathematical model of angiogenic tumor growth in tissue with account of bevacizumab therapy was developed. The model accounts for convective flows that occur in dense tissue under active division of tumor cells, as well as the migration and proliferation dichotomy of malignant cells, which depends on the concentrations of major metabolites, such as oxygen and glucose. Tumor cells wich are in a state of metabolic stress produce vascular endothelial growth factor, which stimulates angiogenesis. To establish the relationship between the capillary network density and oxygen supply, a separate model of stationary blood flow in the capillary network was developed and investigated. A numerical study of the tumor-growth model showed that antiangiogenic bevacizumab treatment of tumors of the diffuse type reduces the total number of their cells, but practically does not affect the rate of their invasion into normal tissues. At the same time, it was found that the growth of dense tumors may be non-monotonic in a rather wide range of parameters. It was shown that in this case bevacizumab therapy stabilizes and significantly inhibits tumor growth, while its local-in-time efficiency is sensitive to the time that it begins.