[A non-invasive method of evaluation of Helicobacter pylori urease activity: its perfection and introduction to clinical practice].

The authors have developed a highly sensitive method of non-invasive diagnostics of Helicobacter pylori, the most frequent human infection. Detection of urease activity is based upon measurement of the degree of the elevation of stable 13C isotope content in exhaled air after administration of C-urea as a test reagent. The method has been scarcely applied in Russia because the test preparation, 13C urea had not been produced domestically until 2002. The method can be easily applied by any healthcare institution; however, it requires special equipment and trained personnel to perform measurement of 13C content in exhaled air samples using mass spectrometers. The article presents the first experience in clinical application of the method.

[A comparative study of the effect of pyrazole on the activity of enzymes in the alcohol/polyol dehydrogenase family]. / Sravnitel'noe issledovanie vliianiia pirazola na aktivnost' fermentov semeistva alkogol'/polioldegidrogenaz.

A comparative study of the effect of pyrazol, an inhibitor of the coenzyme-binding site of alcohol dehydrogenases, on the activity of enzymes of the alcohol/polyol dehydrogenase group has been carried out. Commercial preparations of alcohol dehydrogenases from the cytoplasm of horse liver cells and yeast cells, as well as the enzyme from the cytoplasm of Trichosporon pullulans cells was completely inhibited by 1 mM pyrazol, while alcohol dehydrogenases from Candida utilis and Saccharomyces carlsbergensis were inhibited only by 25% and the enzymes from Saccharomyces cerevisiae and Torulopsis candida by 30 and 38%, respectively. The inhibition degree of alcohol dehydrogenases from the cytoplasm of liver cells of various mammals (bull, calf, rat, gopher) and birds (hen, pheasant, duck) varied from 12 to 42% in the presence of 1 mM pyrazol. The activity of sorbitol dehydrogenase from the liver cytoplasm of these mammals and birds changed neither in the presence of 1 mM pyrazol, nor in the case of a 15-fold increase of the inhibitor concentration. Possible structural differences in the coenzyme-binding site of the active center of the enzymes under study are discussed.

[The possible effect of amino acids forming a loop in the surface layer of subunits on the electrophoretic mobility of enzymes in the alcohol/polyol dehydrogenase family]. / Vozmozhnoe vliianie aminokislot, obrazuiushchikh petliu v poverkhnostnom sloe sub''edinits, na élektroforeticheskuiu podvizhnost' fermentov semeistva alkogol'/polioldegidrogenaz.

Relative electrophoretic mobility (REM) of alcohol dehydrogenases from equine hepatocyte cytoplasm was low probably due to the presence of a loop which consisted of 21 amino acid residues in the surface layer of the enzyme subunits. The REM of multiple molecular forms of alcohol dehydrogenases from yeast cell cytoplasm was higher as consistent with the absence of this loop in the surface layer of the enzyme subunits. Possible role of amino acid residues comprising the loop, in the formation of total charge and their effect on REM values of enzymes from the alcohol/polyol dehydrogenase family are discussed.

[The effect of superoxide dismutase on histochemical detection of dehydrogenases on zymograms using sorbitol dehydrogenase as an example]. / Vliianie superoksiddismutazy na gistokhimicheskoe vyiavlenie degidrogenaz na zimogrammakh na primere sorbitoldegidrogenazy.

Sorbitol dehydrogenases from the cytoplasm of plant, animal, and microbial cells was used to study the effect of superoxide dismutase on histochemical exposure of dehydrogenases after their electrophoretic separation. In some organisms, both enzymes are localized in the same region, which finally leads to the formation of hydrasine tetrazolium, a soluble colourless compound, but not of diformazan, an insoluble stained derivative of tetrazolium nitroblue. The experimental conditions for histochemical exposure of the enzymes in gels are discussed.

[Dot immunoenzyme analysis in biological research]. / Tochechnyi immunofermentnyi analiz v biologicheskikh issledovaniiakh.

The modern state, general methodologic problems and the possibilities to use in biological research the dot enzyme-linked immunosorbent assay (dot-ELISA) are analysed in the review. New types of microporous polymer membranes and equipment for the application of the solid-phase reagent and performing the assay are considered. Different variants of dot-ELISA and methods for the evaluation of results obtained in the assay as well as the ways for its optimization are discussed.

[Activity of alcohol dehydrogenase from cell cytoplasm of several types of yeast]. / Issledovanie aktivnosti alkogol'degidrogenaz iz tsitoplazmy kletok nekotorykh drozhzhei.

The author studied the activity of alcohol dehydrogenase from the cytoplasm of Saccharomyces cerevisiae, Saccharomyces carlsbergensis, Candida utilis and Torulopsis candida cells. The "classical" alcohol dehydrogenase and the octanol dehydrogenase from the T. candida cytoplasm are active in a wide range of pH from 7.0 to 10.5, whereas in the cytoplasm of other yeast cells these enzymes are active in a relatively narrow range of pH. Essential differences were also revealed in the co-enzyme used. In the cytoplasm of S. carlsbergensis and T. candida cells the enzymes display the activity only in the presence of NAD+, while in the other cells, in the presence of both NAD+ and NADP+. The substrate specificity varies as well. While the general activity of alcohol dehydrogenases from the T. candida cytoplasm decreases gradually with the growth of the alcohol radical from C2 to C10, a different regularity is observed in the other cells. Optimal conditions were chosen for maintaining the enzyme activity during prolonged storage of cytoplasm preparations from the yeasts under study.

[The effect of sugars on the activity of sorbitol dehydrogenase from the cytoplasm of bovine liver cells]. / Vliianie sakharov na aktivnost' sorbitoldegidrogenazy iz tsitoplazmy kletok pecheni byka.

The effect of sugar and its phosphate derivatives on sorbitoldehydrogenase from bovine liver has been studied. The presence of 100 mM glucose, mannose, and arabinose did not influence that activity of the studied reaction, whereas fructose, sorbose, and xylose, inhibit the reaction by 20-25%. This can be explained in terms of inhibition by the final reaction products. Inhibition by glucose-6-phosphate (24%), glucose-1-phosphate (21%), and fructose-6-phosphate (42%) is of particular interest since these compounds may play a regulatory role.

[Kinetic properties of sorbitol dehydrogenase from calf liver cell cytoplasm]. / Kineticheskie svoistva sorbitoldegidrogenazy iz tsitoplazmy kletok pecheni telenka.

The kinetic properties of sorbitol dehydrogenase from calf liver cell cytoplasm during sorbitol oxidation were studied at pH 7.0, 7.5, 8.0, 9.0 and 10.0. It was found that the shape of kinetic curves for NADH accumulation depends on pH and substrate concentration. At pH 7.0, 7.5 and 8.0 the enzymatic reaction obeys the Michaelis-Menten kinetics with Km of 3.3 x 10(-3) M. 2.3 x 10(-3) M and 2.08 x 10(-3) M, respectively. At pH 9.0 and 10.0 the vovs [So] curves have an "intermediate plateau". The Hill plots for this reaction reveal two slopes that are dependent on substrate concentration. The nH values for sorbitol (up to 2 mM) are 1.0 and 1.16 at pH 9.0 and 10.0, respectively. With a further rise in the substrate concentration, the nH value increases up to 2.4 and 2.18 at pH 9.0 and 10.0, respectively. This is suggestive of the existence of a slowly dissociating enzymatic system of the Np in equilibrium P type (where P is the oligomeric and p the monomeric forms of the enzyme); N approximately greater than 2. The vovs NAD plots are S-shaped at all pH values studied. The data obtained are discussed in terms of regulatory effects of sorbitol and acidity on association-dissociation of sorbitol dehydrogenase from liver cell cytoplasm.

[The detection of sorbitol dehydrogenase in the cytoplasm of liver cells in birds and its relation to alcohol dehydrogenase and glucose-6-phosphate dehydrogenase]. / Obnaruzhenie sorbitoldegidrogenazy v tsitoplazme kletok pecheni nekotorykh ptits i ee sviaz' s alkogol'degidrogenazoi i gliukozo-6-fosfatdegidrogenazoi.

Comparative studies have been made in the specific activity of sorbitol dehydrogenase, glucose-6-phosphate and alcohol dehydrogenases in the cytoplasm from the liver of wild and domestic ducks, hen and pheasant. High activity of all the three enzymes was found in ducks indicating the effective sorbitol (polyol) metabolism of glucose. The activity of glucose-6-phosphate dehydrogenase is an order lower as compared with the activity of sorbitol and alcohol dehydrogenases in the cytoplasm of hen liver. The same relationship was found for the activity of sorbitol dehydrogenase in the cytoplasm of pheasant liver.

[The rapid electrophoresis of proteins on polyacrylamide gel]. / Ekspress-élektroforez belkov v poliakrilamidnom gele.

A system for express electrophoretic separation of proteins in a thin layer of polyacrylamide gel covalently attached to a glass base has been proposed. A technology of covalent attachment of gel to a glass base has been developed and a small-scale instrument has been constructed which allows to carry out the separation, staining and electrophoregram washing off for 60 min. The quality of the cleaved protein mixtures is not worse than the electrophoregrams obtained using the "Phase System T. M." instrument of "Pharmacia".

[The heterogeneity of sorbitol dehydrogenase from the cytoplasm of mammalian brain cells]. / Geterogennost' sorbitoldegidrogenazy iz tsitoplazmy kletok mozga mlekopitaiushchikh.

The multiple molecular forms of sorbitoldehydrogenase in cytoplasm of brain cells of bull, ground squirrel, guinea-pig, rat, hamster and mouse have been found using the method of electrophoresis in polyacrylamide gel and the subsequent specific dyeing for the fermentative activity. All revealed zones of activity are related to the slowly migrating ones. A set of multiple molecular forms from different sources is various. A form with relative electrophoretic activity 0.385 is found in all analyzed animals. The conditions for obtaining of distinct zones of activity on zymograms are chosen.

[A method of hermetic sealing of glass tubes to be filled with polyacrylamide gel]. / Sposob germetizatsii steklianykh trubok dlia zalivki poliakrilamidnogo gelia.

A reliable and simple system is suggested for hermetic sealing of glass tubes to be filled with polyacrylamide gel. It consists of a disk made of hydrophobic material (teflon, capronyl), whose diameter corresponds to that of the tube and rubber sleeve with an inner diameter of 1 mm less than that of the tube and disk. In assembly the disk closes the bottom end of the tube and is fixed with the rubber sleeve.

[The level of sorbitol dehydrogenase activity in the cytoplasm of mammalian liver cells]. / Uroven' sorbitoldegidrogenaznoi aktivnosti v tsitoplazme kletok pecheni mlekopitaiushchikh.

A comparative study of sorbitol dehydrogenase activity in bovine, calf, and rat liver cell cytoplasm has been carried out. The level of activity of the enzyme is several times greater than that of marker enzymes (alcohol dehydrogenase, glucose-6-phosphate dehydrogenase). The data obtained suggest that the polyol (sorbitol) metabolism pathway of glucose functions actively in mammalian liver cells.

[Conditions for continuous registration of enzyme activity]. / Usloviia nepreryvnoi registratsii fermentativnoi aktivnosti.

Conditions of continuous registration of enzyme activity are considered on the example of alcohol dehydrogenase and sorbitol dehydrogenase from cytoplasm of the bovine liver cells. A device permitting to register the initial steps of enzyme interaction with the effector (substrate, coenzyme or inhibitor) is described. The importance of the reaction product coupling for analysis of enzyme activity is demonstrated.

[Functioning of the sorbitol pathway of glucose metabolism during hibernation]. / Funktsionirovanie sorbitol'nogo puti metabolizma gliukozy v usloviiakh gibernatsii.

Studies have been made on the activity of sorbitol dehydrogenase and glucose-6-phosphate dehydrogenase in the liver of hibernating ground squirrels. It was found that the activity of the former is an order higher than that of the latter. Contribution of sorbitol pathway in total metabolism of the glucose in hibernating ground squirrels is discussed.

[Physico-chemical characteristics of sorbitol dehydrogenase from the cytoplasm of bovine liver cells]. / Fiziko-khimicheskie kharakteristiki sorbitoldegidrogenazy iz tsitoplazmy kletok pecheni byka.

Partially purified preparation of sorbitol dehydrogenase, isolated from hepatocytes of bovine liver tissue, was active at a wide range of pH exhibiting the maximal activity at pH 9.0 in presence of NAD but not of NADP. The high rate of sorbitol and xylitol dehydration was observed, whereas the enzyme dehydrated ribitol at the 4-fold lower rate. Disc electrophoresis of the preparation in polyacrylamide gel, where sorbitol, xylitol and ribitol were used as substrates for colorimetric detection of the enzyme activity, exhibited six enzymatic zones with Rf 0.387, 0.266, 0.338, 0.193, 0.129 and 0.064. Optimal conditions were developed for storage of the active enzyme.

[Physicochemical characteristics of sorbitol dehydrogenase from the cytoplasm of the liver cells of susliks]. / Fiziko-khimicheskie kharakteristiki sorbitoldegidrogenazy iz tsitoplazmy kletok pecheni suslikov.

Some physico-chemical properties of sorbitol dehydrogenase from squirrel liver cell cytoplasm have been investigated. Non-linear dependence of enzyme activity upon media pH is shown. Activity manifests only in the presence of NAD that can't be replaced by NADP. The enzyme exhibits stereospecificity: it dehydrates polyol of any length, the second and the fourth carbon atoms have common L-configuration concerning the first carbon atom. A diffusion zone with Rf = 0,09 is revealed on the electrophoregram. Three thin zones of activity about pH 7,2 are exposed by isoelectrofocusing method.

[Electrophoretic evaluation of multiple molecular forms of enzymes and their ratios using alcohol dehydrogenase from mammalian liver cytosol as an example]. / K voprosu ob élektroforeticheskoi otsenke mnozhestvennykh molekuliarnykh form fermentov i ikh sootnosheniiakh na primere alkogol'degidrogenazy iz tsitozolia pecheni mlekopitaiushchikh.

Zymograms of alcohol dehydrogenase from liver cytosol of horse, rat, pig, bull and calf were studied in order to evaluate the advantages and limitations of polyacrylamide gel disc electrophoresis in estimation of multiple molecular forms of the enzyme and their ratios. The zymograms of these animal species differed in amount of zones of the enzymatic activity, in their relative electrophoretic mobility (Rf) and in intensity of staining. The amount of zones and intensity of staining depended on the time of incubation and pH value. No direct correlation could be found between the period of incubation and intensity of the zones colouring, which was due to a non-linear type of dynamics of the reaction products accumulation. As a precondition for correct evaluation of the zymograms results of enzymological estimations (involving pH optima, temperature, saturating concentrations for substrates and coenzymes) were essential.