[Formation of a new type of dinitrosyl iron complexes bound to cysteine modified with methylglyoxal].

It has been shown that interaction of cysteine dinitrosyl iron complexes with methylglyoxal leads to the formation of a new type of dinitrosyl iron complexes., EPR spectrum of these complexes essentially differs from spectra of dinitrosyl iron complexes containing unmodified thiol. The products of the cysteine reaction with methylglyoxal are hemithioacetals, Schiff bases and thiazolidines, which most likely serve as ligands for the new type of dinitrosyl iron complexes. It has been shown that the new type of dinitrosyl iron complexes as cysteine dinitrosyl iron complexes, which are physiological donors of nitric oxide, exert a vasodilator effect. It has also been found that the oxidative destruction of the new type of dinitrosyl iron complexes occurs at normal oxygen partial pressure, but these dinitrosyl iron complexes remain rather stable under hypoxia modeling. An assumption that the destruction of the new type of dinitrosyl iron complexes is caused by the formation of a bound peroxynitrite-containing intermediate is made.

[Physico-chemical features of dinitrosyl iron complexes with natural thiol-containing ligands underlying biological activities of these complexes].

Current notions and new experimental data of the authors on physico-chemical features of dinitrosyl iron complexes with natural thiol-containing ligands (glutathione or cysteine), underlying the ability of the complexes to act as NO molecule and nitrosonium ion donors, are considered. This ability determines various biological activities of dinitrosyl iron complexes--inducing long-lasting vasodilation and thereby long-lasting hypotension in human and animals, inhibiting pellet aggregation, increasing red blood cell elasticity, thereby stimulating microcirculation, and reducing necrotic zone in animals with myocardial infarction. Moreover, dinitrosyl iron complexes are capable of accelerating skin wound healing, improving the function of penile cavernous tissue, blocking apoptosis development in cell cultures. When decomposed dinitrosyl iron complexes can exert cytotoxic effect that can be used for curing infectious and carcinogenic pathologies.

[Changes in the electrical characteristics of the internal surface of cytoplasmic membrane of bacteria Escherichia coli after the binding of microdoses of copper ions by its external surface].

The effect of microdoses of copper ions bound with the external surface of the cytoplasmic membrane of Escherichia coli spheroplasts on the charge density of its superficial structures was investigated by the ESR method. Positively charged spin probes of KAT(n)-type and the newly synthesized KAT15 were used. The experimental data were analyzed using the formalism of Gouy-Chapman theory. The binding of microdoses of copper ions by the external surface of the cytoplasmic membrane changed the value of charge density on the membrane internal surface.

Isolation and Purification of enzymes from ligninolytic complex of the basidial fungus Trametes pubescens (Schumach.) Pilát and study of their properties.

A method for purification of enzymes from the ligninolityc complex of the basidiomycete Trametes pubescens (Schumach.) Pilat has been elaborated. Two homogeneous isoforms of laccases (laccase 1 and laccase 2) as well as a homogeneous preparation of lignin peroxidase were isolated. Basic biochemical parameters of the enzymes were determined, such as the molecular weights (67, 67, and 45 kD, respectively), isoelectric points (5.3, 5.1, and 4.2, respectively), as well as content and composition of the carbohydrate moiety of the laccases (N-acetylglucosamine, mannose, and xylose). The pH dependences and thermal stabilities of the laccases were investigated. The kinetic parameters of the enzymatic reactions catalyzed by the laccases were determined using different substrates, such as catechol, hydroquinone, 2,2 -azinobis-(3-ethylbenzthiazoline-6-sulfonate), and K4Fe(CN)6. The structure of the active sites of both laccases and the lignin peroxidase were studied by EPR, CD, and UV-VIS spectroscopy, as well as using fluorescence analysis. Our studies showed similarity of the spectral characteristics of the two laccases, whereas their kinetic properties were found to be different.

[Sulfur ligands within the cluster formations of copper at its strong'binding sites on the cytoplasmic membrane of Escherichia coli]. / Sernye ligandy v sostave klasternykh obrazovanii medi v mestakh ee sil'nogo sviazyvaniia tsitoplazmaticheskoi membranoi bakterii Escherichia coli.

The analysis of the saturation curves of ESR signals revealed a decrease in the relaxation rate of fast-relaxation Cu(I) complexes on the cytoplasmic membrane of E. coli after the interaction of these bacteria with low concentrations of SH-reagents. It was concluded that the observed changes are associated with the reorganization of Cu clusters due to the binding of SH groups incorporated into the clusters N-ethylmaleimide or Ag(I).

[Electron spin resonance determination of copper binding site on Escherichia coli cell membrane]. / Issledovanie metodom elektronnogo paramagnitnogo rezonansa tsentrov medi v mestakh ee sil'nogo cviazyvaniia s tsytoplasmatichesko'i membrano'i bakteri'i Escherichia coli.

The electron-spin relaxation of Escherichia coli cytoplasmic membrane strong binding Cu-centers was investigated by means of the microwave power saturation of the electronic spin resonance signal. It has been established, that copper centres in the strong binding sites of the cytoplasmic membrane E.coli may be represented in the following way: 1) isolate copper complexes with small speed of the spin-lattice relaxation; 2) isolate copper complexes with increased speed of spin-lattice relaxation by means of interaction with rapidly relaxation centres; 3) dipol-binding clasters; 4) ESR-nondetectable at T = 40 K, exchange-binding clasters, which cause increasing of the spin-lattice relaxation speed for isolate copper complexes.

[New prospects in assessing the absorbed radiation dose by electron paramagnetic resonance]. / Novye vozmozhnosti v otsenke pogloshcheniia radiatsionnoi dozy metodom élektronnogo paramagnitnogo rezonansa.

Spin-lattice relaxation rates of radiation induced and background centers in dental enamel have been investigated by electron spin resonance techniques. It has been found that these centers differ in the value of the saturation half-power and the saturation process. Based on these experimental results a new approach to discrimination of the radiation induced signal from the total ESR spectrum of dental enamel is proposed. Comparison of the current methods of radiation dose estimation by the ESR analysis has been performed using the new approach.

The source of non-heme iron that binds nitric oxide in cultivated macrophages.

In cultured macrophages (J 774 line) a decrease in iron-sulfur centers (ISC) was not observed after 5 min treatment with nitric oxide (NO) (10(-7) M NO/10(7) cells). The content of these centers was measured by electron spin resonance (ESR) spectroscopy at 16-60 K. However, the appearance of a characteristic ESR signal at g(av) = 2.03 indicated the formation of dinitrosyl iron complex (DNIC) in these cells. These findings suggest that loosely bound non-heme iron (free iron) but not iron from ISC is mainly involved in DNIC formation. ISC might release iron for DNIC formation after their destruction induced by the products of NO oxidation (NO2, N2O3, etc).

[The source of the iron binding with nitric oxide in activated macrophages]. / Ob istochnike zheleza, sviazyvaiushchegosia s oksidom azota v aktivirovannykh makrofagakh.

No decrease in iron-sulphur centers was found in cultured macrophage cells (J774) after the treatment with nitric oxide (10(-7) M NO/10(7) cells) during 5 min. The center content was controlled by the electron spin resonance (ESR) method. The macrophages pretreated with dithionite + methyl viologen showed the formation of dinitrosyl iron complexes (DNIC) with a characteristic ESR signal at g approximately 2.03. The data suggest that loosely bound nonheme iron (free iron) mostly contributes to the formation of these complexes. Iron from iron-containing proteins does not release from these centers under the direct action of nitric oxide. The iron-sulphur centers can be destroyed by the products of nitric oxide oxidation (NO2, N2O3, etc.) as oxidizing and acid agents.

[Iron-sulfur centers in the Staphylococcus aureus respiratory chain]. / Zhelezosernye tsentry v dykhatel'noi tsepi Staphylococcus aureus.

Using low temperature EPR spectroscopy, signals of iron-sulfur centers with g-factors of 2.02 and 1.94 were detected in the respiratory chain of St. aureus membranes. According to their relaxation parameters and redox properties, these iron-sulfur centers are similar to iron-sulfur centers S-1 and S-3 corresponding to succinate dehydrogenases of mitochondria and bacterial membranes.

Several forms of chromaffin granule cytochrome b-561 revealed by EPR spectroscopy.

Low-temperature EPR spectra of chromaffin granule membranes from bovine adrenal medulla reveal 3 different signals of the ferric cytochrome b-561. A typical gZ signal of a low-spin cytochrome observed at g approximately 3 is comprised of a high-potential component with gZ = 3.14 and a low-potential one with gZ = 3.11, the low-potential signal showing significantly faster relaxation. In addition, a highly temperature-sensitive heme signal at g = 3.7 is observed which is fully retained in the preparation of granule membranes with b-561 reduced by 50% but disappears upon full reduction of the cytochrome by ascorbate. The signal is strikingly similar to that of the mitochondrial low-potential cytochrome b heme (bL or b-566). The presence of several forms of b-561 in chromaffin granule membranes may provide a structural basis for the transmembrane electron transfer believe to be catalyzed by this hemoprotein.

[Formation of intermediate products in the reaction of hemoglobin with cyanide]. / Obrazovanie promezhutochnykh produktov v reaktsii gemoglobina s tsianidom.

Interaction between methemoglobin and cyanide was studied by low temperature inhibition method in combination with ESR. A new, not earlier described in literature ESR signal of low spin cyanide complex of this protein was recorded.

[Iron-sulfur centers in the mitochondrial respiratory chain at different stages of cell culture growth of the hamster Cricetulus griceus]. / Issledovanie zhelezosernykh tsentrov dykhatel'noi tsepi mitokhondrii na raznykh stadiiakh rosta kul'tury kletok khomiachka Cricetulus griceus.

The dramatic diminishing of the concentration of the N-2 iron-sulfur centre of NADH-dehydrogenase of mitochondria during the growth of cell culture of hamster fibroblasts with the subsequent recovery of concentration to the initial level was discovered by means of low-temperature ESR spectroscopy. It was concluded that the results obtained are due mainly to the decrease of the number of respiratory chains, but not to the change of the electron transport chain structure.

[Energy transformation in mitochondria: the role of the iron-sulfur center N-2 of NADH-dehydrogenase]. / Transformatsiia énergii v mitokhondriiakh: rol' zhelezosernogo tsentra N-2 NADH-degidrogenazy.

The data on the action of the iron-sulfur centre N-2 in the 1 site of respiration and oxidative phosphorylation coupling obtained by the authors as well as by other investigators are reviewed in terms of conformational - relaxation concept of energy transformation. The study of the direct and reverse electron transport bears testimony to the fact that the N-2 centre with its proper protein surrounding plays the role of the protein transformer of energy in the 1 site.

[The iron-sulfur center N-2 from NADH-dehydrogenase during direct and reverse electron transport in the mitochondria of rat liver and the yeast Endomyces magnusii]. / Issledovanie zhelezosernogo tsentra N-2 NADN-degidrogenazy pri priamom i obratnom transporte élektronov v mitokhondriiakh pecheni krys i drozhzhei Endomyces magnusii.

A dramatic decrease of the rate of transport of reducing equivalents from NADH to coenzyme Q was observed in the 4th metabolic state (by Chance). It was suggested that this decrease is due to the increase of total time of transition from the structural nonequilibrium state to the equilibrium one of the N-2 center. The structural nonequilibrium state of the center N-2 was observed only under the energy-dependent reverse electron transport, when the substrates for the reduction of coenzyme Q were used.

[Study of the iron-sulfur center N-2 from NADH-dehydrogenase in different metabolic states of the mitochondria from liver and heart]. / Issledovanie zhelezosernogo tsentra N-2 NADN-degidrogenazy v razlichnykh metabolicheskikh sostoianiiakh mitokhondrii pecheni i serdtsa.

The structural--equilibrium and nonequilibrium forms of the center N-2 from NADH-dehydrogenase differ in their parametres of the spin-lattice relaxation. The curves of the temperature dependence of the ESR signal intensity become the effective method of the study of the iron-sulphur proteins. The structural nonequilibrium form of the center N-2 was observed in the "4th" metabolic (by Chance) state, but equilibrium form of the center N-2 prevailed in the "3d" state or in the uncoupled state.

[Reverse electron transfer in mitochondria of the yeast Endomyces magnusii grown on sucrose]. / Obratnyi perenos élektronov v mitokhondriiakh drozhzhei Endomyces magnusii, vyrashchennykh na sakharoze.

Some specific features of cytosol-mitochondria interactions during the growth of the Endomyces magnusii yeast on sucrose media were investigated. Under the given experimental conditions exogenous (cytoplasmic) NADH is the main source of reducing equivalents. Using low temperature EPR spectroscopy, it was demonstrated that exogenous NADH can effectively induce reverse electron transfer in the respiratory chain of the yeast. Fluorimetric assays suggest that End. magnusii mitochondria can utilize the reducing equivalents formed by reverse electron transfer for the reductive amination of alpha-ketoglutarate to form glutamate. The intramitochondrial localization of aminotransferases was postulated.