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Applied Biochemistry and Microbiology (v.47, #1)
Microbial biosensors for detection of biological oxygen demand (a Review) by O. N. Ponomareva; V. A. Arlyapov; V. A. Alferov; A. N. Reshetilov (pp. 1-11).
The review briefs recent advances in application of biosensors for determining biological oxygen demand (BOD) in water. Special attention is focused on the principles of operation of microbial BOD sensors; the information about biorecognition elements in such systems and the methods used for immobilization of biological components in film biosensors is summarized. Characteristics of some BOD sensor models are considered in detail.
Hypoxic stress and the transport systems of the peribacteroid membrane of bean root nodules by V. V. Krylova; S. F. Izmailov (pp. 12-17).
When the roots of Vicia faba L. beans were subjected to hypoxic stress, the activity of H+-ATPase on the peribacteroid membrane, as well as the transport of dicarboxylates (malate and succinate) mediated by this enzyme, decreased. Since malate and succinate are the main carbon-containing metabolites involved in the energy supply to bacteroids, this caused a change of the relation type from mutualism to commensalism, and the domination of the eukaryotes over the prokaryotes consequently increased.
Features of Bacillus subtilis IMB B-7023 and its streptomycin-resistant strain by A. A. Roi; I. P. Yatzenko; A. S. Gordienko; I. K. Kurdish (pp. 18-22).
Features of phosphate-mobilizing bacteria Bacillus subtilis IMB B-7023 and its streptomycin-resistant strain were investigated. While cultivated in medium with glucose and glycerophosphate, the growth rate of the antibiotic-marked strain was approximately similar to this parameter for Bacillus subtilis IMB B-7023 but cell sizes were 1.3-fold less. Both strains significantly stimulated the germinating of plant seeds, attached to their roots, and insignificantly differed in antagonistic activity toward phytopathogens and quantitative content of cell fatty acids and phosphatase activity. Streptomycin-resistant strain may be used for monitoring of Bacillus subtilis introduced to agroecosystem.
Photoinduced bactericidal activity of TiO2 films by S. N. Pleskova; I. S. Golubeva; Yu. K. Verevkin; E. A. Pershin; V. N. Burenina; V. V. Korolichin (pp. 23-26).
A decrease in CFU of gram-positive and gram-negative bacteria on the surface of UV illuminated TiO2 films (wavelength of 380 nm) is shown. A 29, 45, and 47% decrease in bacterial viability of Staphylococcus aureus, Staphylococcus epidermidis, and Escherichia coli, respectively, was seen after 12-min exposition. It was first discovered that the reuse of TiO2 films to test a bacterial suspension for viability removes UV-induced bactericidal activity. However, annealing of TiO2 at a temperature above 400°C restores the photoinduced bactericidal activity to its initial state.
pH and oxidation-reduction potential change of environment during growth of lactic acid bacteria: Effects of oxidizers and reducers by D. Soghomonyan; K. Akopyan; A. Trchounian (pp. 27-31).
Decrease of pH and dropping of oxidation-reduction potential have been observed during growing Lactobacillus salivarius 1588 and 3823, Lactobacillus acidophilus 101E, and Lactococcus lactis 3690 in anaerobic conditions in medium with glucose fermentation. These parameters and membrane proton permeability of bacteria ( $$ C_m^{H^ + } $$ ) changed in the mediums with different pH. Oxidizer ferrycianide and reducer DL-dithiothreitol affected the bacterial growth and their changed H+ extrusion from the cells and K+ uptake by the cells in experiment conditions. Application of oxidizers and reducers are suggested for regulation of growth related with H+ ion transport in lactic acid bacteria.
Expression of modified oxidase of D-aminoacids of Trigonopsis variabilis in methylotrophic yeasts Pichia pastoris by V. A. Redo; E. K. Novikova; M. A. Eldarov (pp. 32-37).
Effective recombinant strains Pichia pastoris that produce functionally active hybrid of Trigonopsis variabilis D-aminoacids bond with chitin-connecting domain of chitinase A1 of Bacillus circulans (DAOcbd) were obtained. The dependence of DAOcbd production levels from production of the number of copies of “expression cassette” integrated in the AOX1 locus of recombinant strains was studied. It was indicated that synthesized DAOcbd may be easily purified and immobilized on chitin sorbents and possessed high specific activity. Produced strains and methods of their cultivation and DAOcbd extraction may be used for development of technologies of obtaining of biocatalyzers in technological processes of obtaining of 7-aminocepha-losporane acid.
Production of 4,15-diacetylnivalenol by Fusarium Sambucinum Fuckel var. minus by G. D. Sokolova; V. N. Voznesenskii (pp. 38-41).
Fusarium sambucinum Fuckel var. minus isolate produced unusual for F. sambucinum Fuckel trichothecene metabolite 4,15-diacetylnivalenol (9 mg/l) in conditions of deep cultivation on Myro medium. This compound was identified by TLC, GLC, HPLC, and 1H NMR spectroscopy. Other trichothecenes, 4-acetylnivalenol (3 mg/l) and nivalenol (1 mg/l), were also found in the culture. The observed feature of the studied isolate is assumed to be due to the presence of an additional gene, which encodes cytochrome P450 oxygenase responsible for the introduction of keto group at C-8 and hydroxyl group at C-7 of the trichothecene structure.
Hydroxylation of steroids by Curvularia lunata mycelium in the presence of methyl-β-cyclodextrine by V. A. Andrushina; A. V. Druzhinina; V. V. Yaderets; T. S. Stitsenko; N. E. Voishvillo (pp. 42-48).
Transformation of 16 Δ5-3β-hydroxy- and Δ4-3-ketosteroids of androstane and pregnane classes was carried out using Curvularia lunata mycelium suspended in phosphate buffer with methyl-β-cyclodextrine (MCD). As the result, 20 monohydroxy- and dihydroxy-metabolites, whose structure was determined using spectra of proton magnetic resonance and mass-spectra, have been isolated. Hydroxylation of Δ5-3β-hydroxy-steroids occurred mostly in the C-7α position whereas hydroxylation of Δ4-3-ketosteroids was in the C-11β position. Only androst-4-en-3,17-dione, 9α-hydroxy-androstenedione, and androsta-1,4-diene-3,17-dione were hydroxylated at C-14α position. Besides main 11β-derivatives, the 6β- and 7β-hydroxy-derivatives with yield 10 and 30%, respectively, were isolated during transformation of progesterone and hydroxymethyl pregnadienone. The ratio of MCD to transforming steroid was 1: 1 (mol/mol). Hydrocortisone and 7α-hydroxyandrostenolone with the yield 55 and 77%, respectively, were obtained at the maximal concentrations of cortexolone 20 g/l and androstenolone acetate 10 g/l in the presence of MCD. Absorption of steroids on mycelium, lower speed of their transformation, low concentrations of modifying substrates, and low yield of hydroxyderivatives have been observed in the absence of MCD.
A Novel screening method of cellulase-producing bacteria based on Phytophthora parasitica var. nicotianae by W. Wang; P. Wang; R. Hu (pp. 49-52).
Cellulase is the key to utilize the renewable and abundant cellulose resource, cellulase-producing microorganism is an important source of cellulase. The traditional screening method of cellulase-producing microorganism is low efficacy and not macroscopic. The screening method in this study was based on the interactive culture character between cellulase-producing bacteria and Phytophthora parasitica var. nicotianae on plates, the results indicated that the inhibition zone and cellulase activity of bacterial strains are conformity on the whole, so the screening method is very quickly and apparent
Strain improvement for enhanced production of cellulase in Trichoderma viride by F. Xu; J. Wang; S. Chen; W. Qin; Z. Yu; H. Zhao; X. Xing; H. Li (pp. 53-58).
The filamentous fungi Trichoderma species produce extracellular cellulase. The current study was carried out to obtain an industrial strain with hyperproduction of cellulase. The wild-type strain, Trichoderma viride TL-124, was subjected to successive mutagenic treatments with UV irradiation, low-energy ion beam implantation, atmospheric pressure non-equilibrium discharge plasma (APNEDP), and N-methyl-N′-nitro-N-nitrosoguanidine to generate about 3000 mutants. Among these mutants, T. viride N879 strain exhibited the greatest relevant activity: 2.38-fold filter paper activity and 2.61-fold carboxymethyl cellulase, 2.18-fold β-glucosidase, and 2.27-fold cellobiohydrolase activities, compared with the respective wild-type activities, under solid-state fermentation using the inexpensive raw material wheat straw as a substrate. This work represents the first application of APNEDP in eukaryotic microorganisms.
Decolorization of the anthraquinone dye Cibacron Blue 3G-A with immobilized Coprinus cinereus in fluidized bed bioreactor by A. Moutaouakkil; M. Blaghen (pp. 59-65).
Coprinus cinereus, which was able to decolorize the anthraquinone dye Cibacron Blue 3G-A (CB) enzymatically, was used as a biocatalyst for the decolorization of synthetic solutions containing this reactive dye. Coprinus cinereus was immobilized in both calcium alginate and polyacrylamide gels, and was used for the decolorization of CB from synthetic water by using a fluidized bed bioreactor. The highest specific decolorization rate was obtained when Coprinus cinereus was entrapped in calcium alginate beads, and was of about 3.84 mg g−1 h−1 with a 50% conversion time (t 1/2) of about 2.60 h. Moreover, immobilized fungal biomass in calcium alginate continuously decolorized CB even after 7 repeated experiments without significant loss of activity, while polyacrylamide-immobilized fungal biomass retained only 67% of its original activity. The effects of some physicochemical parameters such as temperature, pH and dye concentration on decolorization performance of isolated fungal strain were also investigated.
The role of laccase and peroxidase of Lentinus (Panus) tigrinus fungus in biodegradation of high phenol concentrations in liquid medium by D. A. Kadimaliev; V. V. Revin; N. A. Atykyan; O. S. Nadezhina; A. A. Parshin (pp. 66-71).
The possibility of the usage of Lentinus tigrinus fungus strain VKM F-3616D for biodegradation of high (up to 5%) phenol concentrations in liquid medium and the involvement of laccase and peroxidase in this process have been studied. L. tigrinus fungus was demonstrated to effectively degrade phenol with easy biomass deletion from the liquid. Decrease in phenol concentration was accompanied by increased secretion level and laccase activity at the preliminary stages of biodegradation, while that of peroxidase was at the latest stages of biodegradation. These enzyme secretions in distinct ratios and consequences are necessary for effective phenol biodegradation. An effective approach for phenol concentration decrease in the waste water of smoking shops in meat-processing factories using L. tigrinus fungus was described.
Enzyme immunassay of alternariol for the assessment of risk of agricultural products contamination by A. A. Burkin; G. P. Kononenko (pp. 72-76).
A highly specific test system based on polyclonal rabbit antibodies targeting a conjugate of alternariol and bovine serum albumin prepared by formaldehyde condensation has been developed for the first time and shown to have the sensitivity of 0.4 ng/ml in the indirect competitive determination of alternariol. The possibility of applying the assay for assessment of the intensity of alternariol contamination of corn, animal feed, and raw materials used for the preparation of the latter has been demonstrated.
Enzyme immunoassay for the determination of hexestrol in meat by M. M. Vdovenko; C. -F. Peng; C. -L. Xu; E. S. Vylegzhanina; A. A. Komarov; I. Yu. Sakharov (pp. 77-81).
An indirect competitive enzyme-linked immunosorbent assay (ELISA) of hexestrol (HES), an anabolic hormone forbidden for use in livestock farming, has been developed. Conditions of ELISA have been optimized by varying the concentrations of the coating conjugate (HES-ovalbumin), anti-HES antiserum, casein, and Tween 20. In the absence of Tween 20 in the reaction mixture, the detection limit (IC10) equaled 0.01 ng/ml, IC50 equaled 0.17 ng/ml, and the working range (IC20–IC80) equaled 0.03–0.86 ng/ml, while, in the presence of 0.05% Tween 20, these values equaled 0.05, 2.9, and 0.26–32.0 ng/ml, respectively. Standard deviation of the analysis results did not exceed 5.4%. If ELISA was performed in the absence of detergents, the recovery value upon HES determination in spiked beef samples ranged from 74 to 147%.
Pectin substances of the callus culture of Silene vulgaris (M.) G. by Ye. A. Günter; Yu. S. Ovodov (pp. 82-86).
Pectin-protein fraction SVC was isolated from the callus culture of the bladder campion (Silene vulgaris). The main components in it were residues of D-galacturonic acid, galactose, arabinose, rhamnose, and protein. Using ion-exchange chromatography, ultrafiltration, and acid and enzymatic hydrolysis, it was shown that SVC contained a mixture of molecules of linear pectin, branched pectin polysaccharide, and pectin-protein polymer. A fragment of the linear chain of galacturonan amounted to more than half of the entire carbohydrate silenan chain. The branched area of the macromolecule is represented by rhamnogalacturonan I. The pectin-protein polymer consisted mainly of protein and weakly branched pectin fragments with molecular mass of more than 300 kDa.
Respiration activity of suspension cell culture of Polyscias filicifolia bailey, Stephania glabra (Roxb.) miers, and Dioscorea deltoidea wall by M. V. Titova; E. A. Berkovich; O. V. Reshetnyak; I. E. Kulichenko; A. V. Oreshnikov; A. M. Nosov (pp. 87-92).
Peculiarities of respiration of cells cultures producing biologically active compounds (isoprenoids and alkaloids) were investigated in order to optimize productivity of culture growth and biosynthesis. It had been revealed that studied cells cultures of Dioscorea deltoidea Wall (producer of furistanol glycosides), Stephania glabra (Roxb.) Miers (producer of stepharin alkaloid) and Polyscias filicifolia Bailey (complex of biologically active agents) differ both in joint respiration activity and in ratio between cytochrome and cyanide-resistant respiration, while changes of rate of total oxygen consumption and activity of alternative oxidase during growth were found to be individual for every investigated culture. Maximum rate of oxygen consumption for cells of D. deltoidea and S. glabra was marked in the period preceding active synthesis of secondary metabolites (lag phase for D. deltoidea and exponential phase for S. glabra). The revealed trends can be used for further monitoring and regulation of growth and biosynthesis of secondary metabolites in producing cell cultures during deep cultivation.
Influence of a hydrothermal treatment on protein fractions of wheat by N. V. Korablyova; T. D. Kasymova (pp. 93-102).
Influence of different temperature modes of a hydrothermal treatment on the protein fractions of wheat, grown in Uzbekistan, has been studied within a temperature range from 40 to 80°C. Using inversed phase and exclusion chromatography, we have revealed that hydrothermal treatment reduces the extract content and causes some changes in the ratio between high- and low-molecular components. If the treatment temperature exceeded 60°C, then, in all cases, except the glutenin fraction, the content of high-molecular components decreased, whereas the content of low-molecular components increased. The glutenin fraction was more subjected to heat influence and demonstrated a higher ability to aggregation, occurring mainly due to the component whose molecular weight was 113.42 kDa. Reduction of the number of free sulfhydryl groups in wheat gluten and its fractions in the case of a temperature increase indicates the oxidation of these groups with formation of new intermolecular disulphide bonds, which, in turn, results in the aggregation of proteins and strengthening of gluten. The obtained results agree with data of our earlier studies of gluten microstructure and fractioning during a hydrothermal treatment.
Chitosan antiviral activity: Dependence on structure and depolymerization method by V. N. Davydova; V. P. Nagorskaya; V. I. Gorbach; A. A. Kalitnik; A. V. Reunov; T. F. Solov’eva; I. M. Ermak (pp. 103-108).
Enzymatic (the action of lysozyme) and chemical (the action of hydrogen peroxide) hydrolysis of chitosans with various degree of acetylation (DA)—25, 17, and 1.5%—was performed. Purification and fractioning of the hydrolysis products were performed using dialysis, ultrafiltration, and gel-penetrating chromatography. Low-molecular (LM) derivatives of the polysaccharide with molecular masses from 17 to 2 kDa were obtained. The study of their antiviral activity against the tobacco mosaic virus (TMV) showed that these samples inhibited the formation of local necroses induced by the virus for 50–90%. The antiviral activity of the LM chitosans significantly increased with the lowering of their polymerization degree. Furthermore, the products of the enzymatic hydrolysis possessed lower activity than the chitosan samples obtained as a result of chemical hydrolysis. It was revealed that the exhibition of the antiviral activity weakly depended on the degree of acetylation of the samples.