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Applied Biochemistry and Microbiology (v.38, #1)
The Antiviral Activity of Chitosan (Review) by S. N. Chirkov (pp. 1-8).
Data on the inhibitory effect of chitosan on viral infections in animals, plants, and microorganisms are reviewed. The effects of the physicochemical parameters and structure of chitosan on its antiviral activity are analyzed. Possible mechanisms of the inhibitory effect of chitosan on viral infections are discussed.
Development of Enzyme Immunoassays for the Herbicide Chlorsulfuron by E. V. Yazynina; A. V. Zherdev; S. A. Eremin; V. A. Popova; B. B. Dzantiev (pp. 9-14).
Antibodies against the herbicide chlorsulfuron have been raised and characterized. Enzyme immunoassays (EIAs) for chlorsulfuron, involving labeled antigen or labeled antibodies, have been developed. The kinetics of antigen–antibody interactions in the EIA systems developed has been studied. Both systems exhibit equal sensitivity (1 ng/ml). The values of the coefficient of variation (CV), determined within the range of chlorsulfuron concentrations to be measured by the systems (1–100 ng/ml), are not in excess of 8%. The possibility of using glucose oxidase as a label in EIAs for chlorsulfuron has been demonstrated. Lack of cross-reactivity with a series of sulfonyl-/arylurea derivatives and triazines makes it possible to recommend the EIA systems developed for chlorsulfuron determination in the environment.
Testing and Isolation of High-Purity Restriction Endonucleases by L. I. Puchkova; T. A. Ushakova; V. K. Mikhailova; G. D. Serov; G. N. Krivopalova; V. E. Repin (pp. 15-19).
A new method of testing restriction nucleases is proposed. This method is based on high-temperature treatment of crude cell extracts. Disrupted cells were heated at 50–60°C, centrifuged, and assayed for restrictases. This method provides the opportunity for screening new enzymes in microbial strains enriched with nonspecific restrictases. High-temperature treatment of cell extracts of certain producers reduces the number of steps of the procedure used for isolating high-purity restrictases; the resulting preparations are capable of maintaining high enzymatic activity during long-term storage. It was shown that high-temperature treatment can be applied not only to thermophilic but also to mesophilic strains of microorganisms of different taxa.
Restriction Endonuclease Sst12I from a Strain of Streptomyces Recognizing the Nucleotide Sequence 5"-CTGCAG-3" by T. A. Ushakova; L. I. Puchkova; V. V. Gutorov; V. N. Kuvshinov; V. E. Repin (pp. 20-22).
A new restriction endonuclease Sst12I belonging to type II and recognizing the sequence 5"-CTGCAG-3" was isolated from the bacterial strain Streptomycessp. St-12. The enzyme hydrolyzes DNA between adenine and guanine residues; thus, it is a true isoschizomer of restrictase PstI. In contrast to PstI, the restriction endonuclease Sst12I hydrolyses DNA both at 37 and 55°C and remains active after long-term storage.
Development of Biosensors for Phenol Determination from Bacteria Found in Petroleum Fields of West Siberia by A. A. Makarenko; I. P. Bezverbnaya; I. A. Kosheleva; T. N. Kuvichkina; P. V. Il'yasov; A. N. Reshetilov (pp. 23-27).
Nine gram-negative bacterial strains, selected from 300 strains isolated from soils of the West Siberian petroliferous basin and growing on oil and oil products, consume phenol as a single carbon and energy source. The strains were used for the development of a sensor bioreceptor. The most active 32-I strain was shown to bear a plasmid responsible for phenol degradation. The plasmid-free derivative of this strain, 32-I-1, did not grow on phenol. The possibility of creating a model biosensor for phenol based on the plasmid-containing 32-I strain is considered. The detection limit for phenol was 5 μM. The optimum conditions for the sensor operation are: pH 7.4, 35°C, and operation time 30 h.
Selection of Ergot Alkaloid Producers by Induced Mutagenesis by I. G. Vepritskaya; L. V. Boichenko; M. U. Arinbasarov; N. F. Zelenkova; N. V. Bobkova (pp. 28-31).
Using the induced mutagenesis technique, a series of genetically modified Clavicepssp. VKM F-2609 strains that display high levels of agroclavine and elymoclavine synthesis were selected by induced mutagenesis. Compared to the parent strain, c106 displayed a 40-fold higher level of agroclavine synthesis, and c66 displayed an eightfold higher level of elymoclavine synthesis. The levels of synthesis of other alkaloids were decreased in these strains. The effects of various carbohydrates on the strain growth and ergot alkaloid biosynthesis was then investigated in both the parent strain and c106. The largest amount of agroclavine was synthesized by c106 strain growing on a medium with maltose.
Factors Contributing to Roquefortine Yield Variability during Cultivation of Penicillium roquefortii by D. M. Boichenko; N. F. Zelenkova; N. G. Vinokurova; B. P. Baskunov (pp. 32-35).
Variability in the roquefortine yield was shown to be associated with its consumption by the mycelium during isolation of the end product, which depended on temperature, time of culture liquid storage, and biomass concentration. This was also related to the presence in chloroform of chlorocarbonic acid ethyl ester that reacted with roquefortine.
Stability of Glucose 6-Phosphate Dehydrogenase Complexed with Its Substrate or Cofactor in Aqueous and Micellar Environment by A. V. Puchkaev; A. P. Vlasov; D. I. Metelitza (pp. 36-44).
Inactivation of glucose 6-phosphate dehydrogenase (G6PDH) complexed with its substrate, glucose 6-phosphate (GP), or cofactor, NADP+, has been studied within the range 20–40°C in three media: (a) 0.04 M NaOH–glycine buffer (pH 9.1); (b) Aerosol OT (AOT) reversed micelles in octane; and (c) Triton X-100 micelles in octane supplemented with 10% hexanol. The enzyme inactivation was characterized quantitatively by first order rate constants, k in(s–1). In the case of G6PDH–NADP+complexes, the values of k inwere independent of the initial concentrations of G6PDH, either in aqueous medium or AOT micelles. The values of k infor the complex G6PDH–GP were inversely related to the initial concentration of the enzyme, in both aqueous and micellar media. When inactivation of both complexes were studied in AOT micelles, minimum values of k incorresponded to the degree of hydration W 0= 16.7; at W 0> 16.7 and W 0< 16.7, k inincreased. Within the range 20–40°C, the values of k inmeasured for both complexes in aqueous medium were significantly lower than those measured in AOT micelles. Temperature dependences of k inwere characterized by inflections in Arrhenius plots, which corresponded, depending on the medium, to certain temperatures from 33.6°C to 40°C. In all media studied, NADP+complexes of the enzyme exhibited higher stability than their GP counterparts. The parameters of G6PDH and G6PDH–NADP+melting, measured by differential scanning microcalorimetry (maximum temperature and half-width of the transition, enthalpy of denaturation, and van't Hoff enthalpy), provided unequivocal evidence of the higher stability of the complex as compared to that of the enzyme. In addition, this approach demonstrated that G6PDH undergoes destabilization in AOT micelles.
Effect of Phytohormones on the Protein-Synthesizing Ability of Rauwolfia serpentinaBenth. Tissue Culture by N. V. Kirillova; V. P. Komov (pp. 45-47).
The accumulation of intracellular protein by the callus culture of Rauwolfia serpentinaBenth. was studied in a standard or phytohormone-containing medium. Changes in the concentration of total protein in cells induced by indolylacetic and naphthylacetic acids were shown to be associated with the effects of these phytohormones on the biosynthesis and degradation of intracellular protein.
Effect of Arsenic on Bacterial Growth and Plasma Membrane ATPase Activity by V. I. Podol'skaya; T. G. Gruzina; Z. R. Ul'berg; A. S. Sokolovskaya; N. I. Grishchenko (pp. 48-52).
The effects of arsenic in the forms of arsenite and arsenate on bacterial growth and plasma membranes' ATPase activity was studied. Correlation of the rate of ATP hydrolysis was found to be correlated with bacterial resistance to toxic arsenic ions. Detoxification of arsenate by resistant cultures of bacteria was suggested to be related to an increase in bacterial ATPase activity and the degree of ATPase mobilization.
Consumption of Organic Carbon Sources and Biosynthesis of Lactic Acid by the Photosynthetic BacteriumRhodobactersp. D-4 by A. Kh. Paronyan (pp. 53-58).
Nonsulfur photosynthetic purple bacteria isolated from the Dzhermuk mineral springs (Armenia) were grown on sugar-containing media and found to be capable of synthesizing L(+)-lactic acid. Various organic compounds were tested as possible sole sources of carbon and an electron donors required to support bacterial growth and biosynthesis of lactic acid under various growth conditions.
Physicochemical Properties of Melanins Produced by the Sterile Form of Inonotus obliquus(“Chagi”) in Natural and Cultivated Fungus by T. A. Kukulyanskaya; N. V. Kurchenko; V. P. Kurchenko; V. G. Babitskaya (pp. 58-61).
Physicochemical properties of pigments isolated from the naturally occurring sterile form of Inonotus obliquus(Fr.) Pil. known as Chagi and comprising the major constituent of the medicine befungin were compared with those of melanins synthesized by this fungus in the culture in order to develop a new medicine. Elemental and functional group analyses, as well as UV-visible, IR, and EPR spectra, and thermolysis studies revealed structural differences in these pigments and allowed for assignment of the naturally produced melanin to allomelanins, whereas that of cultivated fungus was assigned to eumelanins.
Survival of Rhizobiumin a Monoculture and Binary Population with Rhizosphere Bacteria by L. A. Sukhovitskaya; G. V. Safronova; G. M. Klyshko; N. V. Korolenok (pp. 62-67).
The survival of pure cultures of Rhizobium leguminosarumbv. pisumand Rhizobium trifoliiand their interaction with associative diazotrophic and phosphate-mobilizing bacteria after inoculation of sterile soil were studied. The viable heterotypic diazotrophic and rhizobial phosphate-mobilizing association was formed whose efficiency was 14% (clover) and 28% (pea) higher compared to monorhizobial inoculates.
Determination of the Content and Degree of Esterification of Uronic Acids in Plant Tissues and Products of Their Processing by M. P. Filippov; G. V. Chernei (pp. 68-71).
A method for quantitative determination of uronic acids and the degree of esterification of their carboxyl groups in plant tissues and products of their processing is described. The method involves determination of the difference between the concentrations of Cu2+in solution before and after interaction of copper with the substance in question. Copper was determined spectrophotometrically in the form of a copper–ammonium complex.
Isolation and Characterization of Holocellulose from Alfalfa by M. S. Dudkin; E. I. Danilova; L. F. Shchelkunov (pp. 72-77).
Holocellulose isolated from the aerial parts of alfalfa (Medicago sativa) contains a polysaccharide complex of cellulose and hemicelluloses, the major structural components of cell walls. Holocellulose is highly hydrophilic and has a dense biopolymer packing. The carboxylic groups of hemicelluloses and cellulose determines the ability of holocellulose to adsorb polyvalent metal cations.
Volatile Metabolites and External CO2Exchange of Wheat Cenoses under Optimal Conditions and Thermal Stress by I. I. Gitel'son; A. A. Tikhomirov; O. V. Parshina; S. A. Ushakova; G. S. Kalacheva (pp. 78-82).
The effects of elevated temperature (35 and 45°C) on photosynthesis, respiration, and both the qualitative and quantitative compositions of volatile emissions (VE) of wheat (Triticum aestuvumL. cultivar 232) cenoses at light intensities of 70, 150, or 240 W/m2of photosynthetically available radiation (PAR) were studied. At a PAR of 240 W/m2, the thermal stabilities of photosynthesis and respiration increased at 35°C and decreased at 45°C. Elevated temperatures nonuniformly changed the rates and direction of VE syntheses. In this process, the highest increase in VE evolution was observed at 70 W/m2and 35°C; the lowest, at 240 W/m2. In addition, the concentrations and composition of VE during the repair period differed from the initial values.
Autoantibodies to Human Thyroid Peroxidase in Immunoassay and Immunoaffinity Chromatography by I. I. Vashkevich; E. P. Kiseleva; O. V. Sviridov; L. I. Survilo; O. A. Strel'chenok (pp. 83-88).
A biochemical system of radioimmunoassay of human thyroid peroxidase (TPO) with the use of specific autoantibodies was developed for the first time. This system includes lyophilized preparations of human autoantibodies to TPO, radioiodinated TPO, standards prepared from pure TPO, and the solid-phase protein A as a precipitating agent. An analytical system was used to study the TPO distribution in fractions of thyroid tissue homogenate. Differences in TPO concentrations in mitochondria, microsomes, and supernatant fluid depending on thyroid pathology and tissue storage conditions were found. The behavior of immobilized anti TPO autoantibodies in immunoaffinity chromatography of this microsomal antigen was studied.
An Insoluble Colored Substrate for Dextranase Assay by E. F. Khalikova; N. G. Usanov (pp. 89-93).
An assay of dextranase (EC 3.2.1.11) was developed by using Sephadex G-200 coupled with Remazol Brilliant Blue (RBB) as an insoluble substrate. The assay procedure included incubation of the suspension of the colored substrate in a buffer containing the enzyme under study, removal of a residual insoluble substrate, and measurement of the absorbance of supernatant fluid containing colored soluble hydrolysis products at 595 nm. The procedure was examined in the screening of dextranase-forming bacilli from the microbial collection of the Institute of Biology, Ufa Research Center, RAS.