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Applied Biochemistry and Microbiology (v.45, #4)
The protective action of cytokinins on the photosynthetic machinery and productivity of plants under stress (review) by I. I. Chernyad’ev (pp. 351-362).
A putative way of the protective action of cytokinins on the photosynthetic processes in crops experiencing various stress factors is considered. Various cytokinins are characterized. Pathways of the multiple effects of cytokinin preparations mediating the protection of the photosynthetic machinery from stress are described. Cytokinins interact with receptor proteins, and then the signal is transduced to primary cellular targets (primary response genes). These genes, which possess receptor domains, induce synthesis of the corresponding mRNAs and photosynthesis-related proteins of chlorophyll-protein complexes, the electron-transport chain, and carbon metabolism, primarily, the key enzyme ribulose bisphosphate carboxylase/oxygenase. The protective action of cytokinins under stress conditions preserves the structure and function of the photosynthetic machinery. The application of cytokinins to improving crop yields is discussed.
The purification and characterization of a novel lipid transfer protein from caryopsis of barnyard grass (Echinochloa crusgalli) by E. A. Rogozhin; T. I. Odintsova; A. Kh. Musolyamov; A. N. Smirnov; A. V. Babakov; Ts. A. Egorov; E. V. Grishin (pp. 363-368).
A novel lipid transfer protein called Ec-LTP was isolated from resting caryopsis of weed barnyard grass Echinochloa crusgalli (L.) Beauv.; its molecular weight, amino acid content and N-terminal amino acid sequence were determined. Ec-LTP was a 9150 D protein, containing eight cysteine residues, which formed four disulfide bonds. The isolated protein could significantly inhibit the development of pathogenic fungi Phytophthora infestans and Helminthosporium sativum, causing the late blight of potato and tomato and the root rot of herbs, respectively.
Ribonuclease from the hepatopancreas of the red king crab Paralithodes camtschatica by N. I. Menzorova; Ju. T. Sibirtsev; V. A. Rasskazov (pp. 369-373).
Three enzyme preparations, two acid and one alkaline RNases, were isolated from the hepatopancreas of the red king crab Paralithodes camtschatica using DEAE-cellulose chromatography and gel chromatography. The alkaline RNase was activated by Mg2+ ions and had a pH optimum of 7.2; the acid RNases, a pH optimum of 5.5. The molecular weight of the alkaline RNase was 19 kDa; two acid RNases, 33 and 70 kDa, respectively. The enzymes exhibited a sufficiently high thermostability (IT50 = 53–55°C) and were strongly inhibited by NaCl (IC50, 0.1–0.25 M). The alkaline RNase exhibited no specificity for heterocyclic bases, whereas the acid RNases hydrolyzed poly(U) and poly(A) at maximum rates.
The use of enzymatic preparation for the production of low molecular-weight chitosan from the king crab hepatopancrease by A. V. Ilyina; D. S. Zagorskaya; A. N. Levov; A. I. Albulov; N. P. Kovacheva; V. P. Varlamov (pp. 374-379).
This article describes the optimal conditions for the enzymatic hydrolysis of chitosan and its chemically-modified derivatives using the preparation extracted from the king crab hepatopancrease possessing pronounced hydrolythic activity. The following preparations were used: chitosan with a molecular weight of 700 kDa and an acetylation level of 0.15, carboxymethyl chitosan 200 kDa witih an extent of replacement of 0.23, and N-succinyl chitosan 390 kDa with an extent of replacement of 0.8. Low molecular-weight samples of chitosan and of its modified derivatives were obtained with the yields of 85, 55, and 80%, respectively. The conditions of the hydrolysis were as follows: an enzyme: substrate ratio of 1: 200, 37°C, and 20 h duration of hydrolysis.
Macromolecular systems and bactericidal films based on chitin derivatives and silver nanoparticles by L. N. Shirokova; V. A. Alexandrova; E. M. Egorova; G. A. Vihoreva (pp. 380-383).
A new polymer composite based on carboxymethylchitin and silver nanoparticles was obtained in order to produce biodegradable wound coating films. The number of metal nanoparticles in the composite may be easily regulated as was verified by UV-VIS-spectroscopy data. A comparative evaluation of silver nanoparticle size in the initial system and in the polymer composition was performed by means of photon correlation spectroscopy. Composite films revealed a pronounced concentration-dependent antibacterial activity towards strains Salmonella typhimurium and Staphilococcus aureus.
Microbial polymers as a degradable carrier for pesticide delivery by O. N. Voinova; G. S. Kalacheva; I. D. Grodnitskaya; T. G. Volova (pp. 384-388).
The possibility of use of polyhydroxyalkanoates (PHAs), biodegradable microbial polyesters, as a carrier for pesticides (α-hexachlorcyclohexane and lindane) for targeted and controlled delivery of these compounds to soil was investigated. The kinetics of polymer degradation and the dynamics of pesticide release from the extended-release formulations was studied. It is shown that pesticides embedded in a degradable polymer (PHA) carrier are released gradually and slowly, without surges, as the polymer is degraded by the soil micro-flora. The microbial soil component actively responded to the addition of the polymer as an additional nutrient substrate: the latter was degraded and then utilized. The rate of the pesticide release to the soil can be regulated by varying the polymer-pesticide ratio.
Catalytical properties of Arthrobacter nicotianae cells, a producer of glucose isomerase, immobilized inside xerogel of silicium dioxide by L. V. Perminova; G. A. Kovalenko; N. A. Rudina; L. I. Sapunova; I. O. Tamkovic; A. G. Lobanok (pp. 389-394).
Arthrobacter nicotianae cells, producers of glucose isomerase, were immobilized inside xerogel of silicium dioxide, and properties of the resulted heterogeneous biocatalysts were investigated in the process of isomerization of monosaccharide (glucose and fructose). The glucose isomerase activity of the resulted biocatalysts was shown to be 10 U/g, on average, taking into account the loss of the activity upon the immobilization, which amounted to 50% of the cell activity in suspension. The rate of the fructose isomerization increased linearly in the range of 55–80°C with the temperature coefficient 1.3. The biocatalysts were stable in this range; they were rapidly inactivated, however, at increasing temperature. The half-pife time of inactivation was six to seven h and five min or less at 80 and 85°C, respectively. The half-pife time of inactivation of heterogeneous biocatalysts was 50–90 h in the periodic process of isomerization of 2 M monosaccharides at 60°C in the presence of the immobilized Arthrobacter nicotianae cells.
The introduction of the 9α-hydroxy group into androst-4-en-3,17-dione using a new actinobacterium strain by N. V. Rodina; V. A. Andryushina; T. S. Stytsenko; T. P. Turova; R. V. Baslerov; A. N. Panteleeva; N. E. Voishvillo (pp. 395-400).
A 9α-hydrolase activity of a new actinobacterium strain identified as Rhodococcus erythropolis based on the analysis of a 16S rRNA gene sequence (1417 nucleotides) towards androst-4-en-3,17-dione (AD) was studied. In the presence of glucose in the medium, this strain completely transformed AD (4–20 g/l) into 9α-hydroxy-AD over 20–48 h. This culture was able to grow and perform AD 9α-hydroxylation at a concentration of dimethyl formamide up to 9%. Crystalline 9α-hydroxy-AD was isolated with a yield of over 90%.
Periodic bioleaching of refractory gold-bearing pyrite ore by N. S. Vardanyan; S. Z. Nagdalyan (pp. 401-405).
The main characteristics of a periodic bioleaching of the refractory gold-bearing pyrite ore from the Tandzut deposit (Armenia) with the help of moderate thermophilic bacterium Sulfobacillus thermosulfidooxidans subsp. asporogenes and original thermotolerant strains Leptospirillum spp. were studied. The optimal pH for oxidizing the ore by S. thermosulfidooxidans subsp. asporogenes was 1.8; the pulp density providing maximal iron leaching rate was 10%. The intensity of oxidation processes decreased at higher ore concentrations. When using S. thermosulfidooxidans subsp. asporogenes, the largest amount of iron passed into the solution at the initial oxidant (Fe3+) concentration of 1.3 g/l. Cocultivation of S. thermosulfidooxidans subsp. asporogenes and Leptospirillum spp. increased the degree of pyrite ore leaching to 98.4% vs. 34.1% in the case of the former bacterium alone.
Oxidation of sulfur-containing substrates by an association of acidophilic chemolithotrophic microorganisms by A. A. Revenko; V. S. Melamud; G. S. Krasil’nikov; T. A. Pivovarova; A. V. Belyi; T. F. Kondrat’eva (pp. 406-413).
Quantitative and qualitative changes in the content of elements in the solid and liquid phases occurred as the pulp moved through reactors during biooxidation of an ore flotation concentrate. The association of microorganisms were adapted for utilizing sulfur-containing substrates; however, the rate of their oxidation was insufficient, which led to an increase in the amount of sodium cyanide required for gold recovery. The replacement of one-fourth of the liquid phase of the pulp (density, 13%) with a mineral medium without an energy source, the fractional addition of FeSO4 · 7H2O (1 g/l per day), and the improvement of pulp aeration made it possible to increase the content of SO 4 2− by 80.7, 86.2, and 58.5%, respectively. When one-fourth of the liquid phase of the pulp (density, 24%) was replaced with a mineral medium without an energy source, the rate of additional oxidation of sulfide minerals increased, which increased the efficiency of gold extraction into solution and gold recovery on charcoal by 3.4 and 3.6%, respectively, and reduced sodium cyanide consumption by 3 kg/ton.
Lactobacilli and dairy propionibacterium with potential as biopreservatives against food fungi and yeast contamination by P. -H. Ho; J. B. Luo; M. C. Adams (pp. 414-418).
Naturally fermented and raw foods contain a range of organisms that may have benefit as additives in some foods and food processing. In particular, potential antifungal properties of these organisms may be potentially utilised as natural alternatives to chemical additives used to delay and prevent spoilage by fungi and yeast. This study examined 12 novel bacteria previously isolated from food as possible biopreservatives. The bacteria from the lactobacilli and dairy propionibacterium groups were tested by agar overlay method for their ability to inhibit the growth of 10 fungi and one yeast commonly associated with food contamination. Eight among eleven tested lactic acid bacteria demonstrated broad spectrum of antifungal activity. Strong fungi inhibition was also demonstrated by the dairy propionibacterium, but efficacy was growth medium dependant. Only one fungi, Geotrichum candidum was highly resistant to the bacteria. Variation between the inhibition results for different bacteria identifies the importance of careful strain selection, and the benefits of strain combinations when selecting biopreservatives for foods.
Antifungal and phytostimulating characteristics of Bacillus subtilis Ch-13 rhizospheric strain, producer of bioprepations by V. K. Chebotar’; N. M. Makarova; A. I. Shaposhnikov; L. V. Kravchenko (pp. 419-423).
Bacillus subtilis Ch-13 industrial strain was shown to have a wide spectrum of antagonistic activities against different species of phytopathogenic fungi and bacteria. The B. subtilis Ch-13 strain produces lytic enzymes; cyanide and other antifungal metabolites; stimulates plant growth, producing phytohormones—auxin derivatives. This strain by 2.5 times reduced the quantity of tomato plants infected with phytopathogenic fungus Fusarium oxysporum during inoculation. Fungi abundance on roots with bacterial inoculation was 6.9 times less than in the absence of inoculation. The application of detected antifungal metabolites as biochemical markers for the strain enables to control the stability of physiologic and biochemical characteristics of the producer, and ensures a rapid quality assay of biopreparations with high performance liquid chromatography (HPLC).
The effects of ultraviolet irradiation upon structure and antioxidant activity of silenan from bladder campion callus by E. A. Günter; M. F. Borisenkov; Yu. S. Ovodov (pp. 424-428).
Effects of UV-B (280–315 nm) and UV-C (254 nm) at various doses upon callus of bladder campion (Silene vulgaris (M.) G. were studied. It was revealed that UV irradiation results in the decrease in arabinose and galactose residues in the silenan—the pectin fraction isolated from callus. The silenan possesses antioxidant activity (AOA) as assessed by the reaction with a stable radical. At the irradiation of callus by UV, the AOA of the silenan and the relative content of phenolic compounds in it increased; the highest increase was observed after the irradiation of callus by UV-B. Positive correlation between the AOA of the pectin fraction and an increase in phenolic compounds was revealed. This evidences that the AOA of the silenan relates to and is partially determined by phenolic compounds in its composition. The UV irradiation may be used as a tool to modify the structural features of the cell walls’ polysaccharides in order to produce physiologically-active polysaccharides with desired properties.
The inhibition of potato tubers wound reparation by G. I. Chalenko; N. I. Vasyukova; N. G. Gerasimova; O. L. Ozeretskovskaya (pp. 429-432).
The multiple washing of the wound surface of potato tubers by water adversely affected the protective properties of wound periderm. Immune inhibitor β-1,3-β-1,6 glucan had a property of local effect and inhibited the process of wound healing. The pentasaccharide of xyloglucan caused necrosis of potato tuber tissue and prevented the wound reparation process.
Hydrogen peroxide production in wheat leaves infected with the fungus Septoria nodorum Berk. Strains with different virulence by I. V. Maksimov; A. Sh. Valeev; E. A. Cherepanova; L. G. Yarullina (pp. 433-438).
The effect of two strains of the phytopathogenic fungus Septoria nodorum Berk. of different virulence on the intensity of local generation of hydrogen peroxide in common wheat leaves and the role of oxidoreductases in this process was studied. Differences in the pattern of hydrogen peroxide production in wheat plants infected with high- and low-virulence pathogen strains have been found. The low-virulent S. nodorum strain caused a long-term hydrogen peroxide (H2O2) generation in the infection zone, whereas the inoculation of leaves with the highly virulent strain resulted in a transient short-term increase in the H2O2 concentration at the initial moment of contact between the plant and the fungus. It was shown that the low level of H2O2 production by plant cells at the initial stages of pathogenesis facilitates S. nodorum growth and development. The decrease in the H2O2 concentration induced by the highly virulent S. nodorum strain is determined by inhibition of the oxalate oxidase activity in plant tissues and by the ability of the fungus to actively synthesize an extracellular catalase. The pattern of hydrogen peroxide generation at the initial stages of septoriosis may serve as an index of virulence of S. nodorum population.
Design of experimental approaches on the base of standard proteins for testing blood biopreparations and immunoperoxidase conjugates specific to human and animal immunoglobulines G by A. K. Barsukov; A. V. Barmin; A. I. Kuznetsov; O. Yu. Nesterova; S. A. Ushnurtseva; A. N. Panin; V. I. Smolenskiy; V. I. Ulasov; V. L. Sviderskiy; A. E. Hovanskih (pp. 439-443).
Using standard forms of immunoglobulin (Ig) G and albumin, we have studied electrophoretic and chromatographic profiles of samples of pharmaceutical blood biopreparation batches. The usability of standard proteins was also demonstrated by testing analytical characteristics of immunoperoxidase conjugates specific to human and animal IgG (anti-IgG IPC). In particular, we suggest an additional estimation of analytical characteristics of anti-IgG IPC by the enzyme reaction kinetics with the standard dilution which is calculated by the direct enzyme-liked immunoassay on the homologous IgG-antigen.
Estimation of methods of protein delivery into mammalian cells — A comparative study by electroporation and Bioporter assay by R. Todorova (pp. 444-448).
To test the efficiency of some alternative methods for protein delivery into live cells were used rEGFP (recombinant Enhanced Green Fluorescent Protein) and HeLa cells as model system. The compared methods, electroporation (up to 10%) and Bioporter (up to 20%), deliver proteins and peptides to the cells with efficiency, enough for functional studies of mammalian cells. Thus, the delivery by either of both methods, followed by fluorescence activated cell sorting (FACS) of the cell population, can give a sufficient number of uniformly loaded cells for different studies including transcription assays and gene therapy.
Evaluation of the activity of hydrolases and their inhibitors using substrates immobilized on agarose gel by I. A. Shpirnaya; I. A. Umarov; N. D. Shevchenko; R. I. Ibragimov (pp. 449-453).
A simple and convenient method for the quantitative evaluation of the activity of hydrolytic enzymes and their inhibitors has been proposed. This method is based on the immobilization of a substrate on agarose gel and the following evaluation of the enzyme activity by measuring the hydrolysis area around the well containing the enzyme solution. The method was shown to be applied to the evaluation of the activity of proteinases and amylases and the inhibitors of these enzymes in different biological objects.
Gibberellin A3 detection in plants with high-performance liquid chromatography by E. E. Nefed’eva; N. G. Mazey (pp. 454-458).
A new method of gibberellin A3 detection based on high-performance liquid chromatography has been elaborated. It combines high accuracy and low cost, and includes the following stages: fixation with liquid nitrogen; extraction of alcohol-soluble substances; purification with freezing, reextraction in ethyl acetate, and thin-layer chromatography; and detection with HPLC. Chromatographic conditions and methods of substance identification have been determined. The method allows the use of modest amounts of plant material (5–20 g), at which variability does not exceed 7%, and the total loss of substance is approximately 3%. The results of gibberellin A3 detection in buckwheat are presented.
Detection of polymeric forms of dihydroquercetin by optical absorption and light scattering by N. L. Vekshin (pp. 459-462).
Two methods for the detection of long polymers in dihydroquercetin (DHQ) preparations has been developed. The first method is based on UV spectrophotometry. It was shown that the quantity of long polymers in aqueous solutions can be estimated by the ratio of the absorption bands at 328 and 290 nm, since the 328-nm band was attributed to the monomeric form of DHQ, whereas the 290-nm band was attributed to both the monomeric and polymeric forms. The second method is based on the high-sensitive measurement of light-scattering intensity in aqueous solutions of diluted DHQ preparations using a spectrofluorometer with crossed monochromators. It has been shown that the filtration of DHQ solutions through Millipore filters with a pore diameter of 0.05–0.45 microns makes it possible to nearly completely eliminate long polymers and their aggregates. Long polymers at high concentrations can aggregate. The longest polymers and their aggregates may be 0.1 mm in length, which leads to fluctuations in the light-scattering intensity on the second and minute time scale.