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Individual stages of bacterial dichloromethane degradation mapped by carbon and chlorine stable isotope analysis

Journal of Environmental Sciences 2019年第4期31卷 147-160页

作者： Maria L.Torgonskaya Anatoly M.Zyakun Yuri A.Trotsenko Kestutis S.Laurinavichius Steffen Kümmel Stéphane Vuilleumier Hans H.Richnow Laboratory of Radioactive Isotopes G.K.Skryabin Institute of Biochemistry and Physiology of MicroorganismsFRC Pushchino Center for Biological ResearchRussian Academy of Sciences Laboratory of Mass Spectrometry G.K.Skryabin Institute of Biochemistry and Physiology of MicroorganismsFRC Pushchino Center for Biological ResearchRussian Academy of Sciences Department of Isotope Biogeochemistry Helmholtz Centre for Environmental Research-UFZ Université de Strasbourg CNRS

The fractionation of carbon and chlorine stable isotopes of dichloromethane(CH_2Cl_2,DCM)upon dechlorination by cells of the aerobic methylotroph Methylobacterium extorquens DM4 and by purified DCM dehalogenases of the glutathione S-transferase family was *** effects for individual steps of the multi-stage DCM degradation process,including transfer across the cell wall from the aqueous medium to the cell cytoplasm,dehalogenase binding,and catalytic reaction,were *** observed carbon and chlorine isotope fractionation accompanying DCM consumption by cell supensions and enzymes was mainly determined by the breaking of C\Cl bonds,and not by inflow of DCM into *** isotope effects of DCM dehalogenation were initially masked in high density cultures,presumably due to inverse isotope effects of non-specific DCM oxidation under conditions of oxygen *** cofactor supply remarkably affected the correlation of variations of DCM carbon and chlorine stable isotopes(Δδ^(13)C/Δδ^(37)Cl),increasing corresponding ratio from 7.2–8.6 to 9.6–10.5 under conditions of glutathione *** suggests that enzymatic reaction of DCM with glutathione thiolate may involve stepwise breaking and making of bonds with the carbon atom of DCM,unlike the uncatalyzed reaction,which is a one-stage process,as shown by quantum-chemical modeling.

关键词： Dichloromethane(DCM) Microbial degradation Aerobic methylotrophic bacteria Dichloromethane dehalogenase Carbon and chlorine isotopic fractionation

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Physiological and biochemical characteristic of Miscanthus×giganteus grown in heavy metal-oil sludge co-contaminated soil

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Journal of Environmental Sciences 2022年第5期34卷 114-125页

作者： Anna Muratova Yelena Lyubun Irina Sungurtseva Olga Turkovskaya Asil Nurzhanova institute of biochemistry and physiology of Plants and microorganisms RAS Saratov 410015Russia institute of Plant Biology and Biotechnology Almaty 050040Kazakhstan

The effect of oil sludge and zinc,present in soil both separately and as a mixture on the physiological and biochemical parameters of Miscanthus×giganteus plant was examined in a pot *** opposite effect of pollutants on the accumulation of plant biomass was established:in comparison with uncontaminated control the oil sludge increased,and Zn reduced the root and shoot *** sludge had an inhibitory effect on the plant photosynthetic apparatus,which intensified in the presence of *** specific antioxidant response of M.×giganteus to the presence of both pollutants was a marked increase in the activity of superoxide dismutase(mostly owing to oil sludge)and glutathione-S-transferase(mostly owing to zinc)in the *** participation of glutathione-S-transferase in the detoxification of both the organic and the inorganic pollutants was *** inhibited the activity of laccase-like oxidase,whereas oil sludge promoted laccase and ascorbate oxi-dase *** finding suggests that these enzymes play a part in the oxidative detox-ification of the organic *** both pollutants used jointly,Zn accumulation in the roots increased 6-fold,leading to increase in the efficiency of soil clean-up from the *** turn,Zn did not significantly affect the soil clean-up from oil *** study shows for the first time the effect of co-contamination of soil with oil sludge and Zn on the phys-iological and biochemical characteristics of the bioenergetic plant M.×*** data obtained are important for understanding the mechanisms of phytoremediation with this plant.

关键词： Miscanthus×giganteus Oil sludge Zinc Antioxidant enzymes Oxidative enzymes

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Image Analysis in Microbiology: A Review

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Journal of Computer and Communications 2016年第15期4卷 8-32页

作者： Evgeny Puchkov All-Russian Collection of microorganisms Skryabin Institute of Biochemistry and Physiology of Microorganisms RAS Pushchino Russia

This review is focused on using computer image analysis as a means of objective and quantitative characterizing optical images of the macroscopic (e.g. microbial colonies) and the microscopic (e.g. single cell) objects in the microbiological research. This is the way of making many visual inspection assays more objective and less time and labor consuming. Also, it can provide new visually inaccessible information on relation between some optical parameters and various biological features of the microbial cul-tures. Of special interest is application of image analysis in fluorescence microscopy as it opens new ways of using fluorescence based methodology for single microbial cell studies. Examples of using image analysis in the studies of both the macroscopic and the microscopic microbiological objects obtained by various imaging techniques are presented and discussed.

关键词： Computer Image Analysis microorganisms Viability Yeast Bacteria Fungi Colony Counter Microbial Identification Multispectral Imaging Hyperspectral Imaging Diffraction Pattern Imaging Scatter Pattern Imaging Multifractal Analysis Support Vector Machines Principal Component Analysis Linear Discriminant Analysi ImageJ Matlab Fluorescence Microscopy Microfluorimetry Green Fluorescent Protein (GFP)

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Surface-enhanced Raman scattering inside Au@Ag core/shell nanorods

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Nano Research 2016年第8期9卷 2303-2318页

作者： Boris Khlebtsov institute of biochemistry and physiology of Plants and microorganisms Russian Academy of Sciences Saratov 410049 Russia Saratov National Research State University Saratov 410012 Russia

The design and synthesis of plasmonic nanoparticles with Raman-active molecules embedded inside them are of significant interest for sensing and imaging applications. However, direct synthesis of such nanostructures with controllable shape, size, and plasmonic properties remains extremely challenging. Here we report on the preparation of uniform Au@Ag core/sheU nanorods with controllable Ag shells of 1 to 25 nm in thickness. 1,4-Aminothiophenol （4-ATP） molecules, used as the Raman reporters, were located between the Au core and the Ag shell. Successful embedding of reporter molecules inside the core/shell nanoparticles was confirmed by the absence of selective oxidation of the amino groups, as measured by Raman spectroscopy. The dependence of Raman intensity on the location of the reporter molecules in the inside and outside of the nanorods was studied. The molecules in the interior showed strong and uniform Raman intensity, at least an order of magnitude higher than that of the molecules on the nanoparticle surface. In contrast to the usual surface-functionalized Raman tags, aggregation and clustering of nanoparticles with embedded molecules decreased the surface-enhanced Raman scattering （SERS） signal. The findings from this study provide the basis for a novel detection technique of low analyte concentration utilizing the high SERS response of molecules inside the core/shell metal nanostructures. As an example, we show robust SERS detection of thiram fungicide as low as 10-9 M in solutions.

关键词： surface-enhanced Ramanscattering (SERS) Au@Ag nanorods nanocuboids multipole plasmons

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Polyphosphatase PPX1 of <i>Saccharomyces cerevisiae</i>as a Tool for Polyphosphate Assay

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Advances in Enzyme Research 2015年第4期3卷 93-100页

作者： Lidiya Lichko Tatiana Kulakovskaya skryabin institute of biochemistry and physiology of microorganisms Russian Academy of Sciences Pushchino Russia

The recombinant exopolyphosphatase PPX1 with a specific activity of ~300 U/mg protein was purified from the strain of Saccharomyces cerevisiae with the inactivated PPN1 gene transformed by the expression vector carrying the yeast PPX1 gene. The recombinant PPX1 was similar to the PPX1 of wild strains in its substrate specificity and requirement for cations. PPX1 had the high substrate specificity to polyphosphates. The preparation was suitable for polyphosphate assay in the presence of orthophosphate and nucleoside phosphates not hydrolyzed by PPX1. The yield of the enzyme preparation was 250 assays per 1 g of the biomass. The recombinant PPX1 was successfully used in polyphosphate assay in different yeast species and some foodstuffs.

关键词： Saccharomyces cerevisiae PPX1 Polyphosphate Polyphosphatase Assay Food

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SERS-based lateral flow immunoassay of troponin I by using gapenhanced Raman tags

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Nano Research 2019年第2期12卷 413-420页

作者： Boris N. Khlebtsov Daniil N. Bratashov Nadezhda A. Byzova Boris B. Dzantiev Nikolai G. Khlebtsov institute of biochemistry and physiology of Plants and microorganisms Russian Academy of Sciences Saratov 410049 Russia Saratov State University Saratov 410012 Russia A. N. Bach institute of biochemistry Research Centre of Biotechnology Russian Academy of Sciences Moscow 119071 Russia

The lateral flow immuno assay (LFIA) has emerged as a powerful tool for rapid scree ning owing to its simplicity and flexibility for detection of various biomarkers. However, conventional LFIA strips have several disadvantages, including limits in quantitative analysis and low sensitivity. Here we developed a novel surface-e nhanced Rama n scatteri ng LFIA based on non spherical gap-e nhanced Raman tags (GERTs), with Raman molecules (RMs) embedded in a 1-nm gap between Au nanorod core and Au shell. Such tags have a strong and uniform surface-enhanced Rama n scattering (SERS) resp on se, an order of mag nitude higher than that of other comm on SERS tags such as Au nano rods, nano stars, Au nano shells with surface-adsorbed RMs, or spherical GERTs with embedded RMs. The feasibility of the tags was dem on strated by the semiqua ntitative and sen sitive detecti on of the heart disease biomarker cardiac tropo nin I (cTnI). GERTs were conjugated with mono clonal antibodies and used for LFIA in the same way as ordinary functionalized colloidal gold. The presenee of the target antigen, cTnI, was identified by Raman microscopy mapping of the test zone. With the SERS-based LFIA, the limit of cTnI detection was about 0.1 ng/mL. This value is within the diagnostic range of cTnI in the blood serum of patients with heart infarction and is 30 times lower than that of the colorimetric LFIA test using the same antibodies and either GERTs or colloidal gold as labels.

关键词： surface-enhanced Raman scattering (SERS) lateral flow immunoassay Au core/shell nanorods gap-enhanced Raman tags cardiac troponin I

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Computer Image Analysis as a Tool for Microbial Viability Assessment: Examples of Use and Prospects

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Journal of Biosciences and Medicines 2014年第3期2卷 1-6页

作者： Evgeny Puchkov All-Russian Collection of microorganisms Skryabin Institute of Biochemistry and Physiolog of Microorganisms Pushchino Russia

Application of the computer image analysis for improving microbial viability assessment by plate count and fluorescence microscopy was investigated. Yeast cells were used as a model microorganism. The application of the improved methods for the viability assessment of yeast cells after preservation by freezing and freeze-drying was demonstrated.

关键词： Microbial Viability Microbial Preservation Plate Count Yeast Computer Image Analysis Fluorescence Microscopy Saccharomyces cerevisiae Cryptococcus terreus Xanthophyllomyces dendrorhous

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Au-nanocluster-loaded human serum albumin nanoparticles with enhanced cellular uptake for fluorescent imaging

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Journal of Innovative Optical Health Sciences 2016年第2期9卷 17-24页

作者： Boris Khlebtsov Artur Prilepskii Maria Lomova Nikolai Khlebtsov institute of biochemistry and physiology of Plants and microorganisms Russian Academy of Sciences 13 Prospekt Entuziastov Saratov 410049 Russia Saratov State University 83 Ulitsa Astrakhanskaya Saratov 410012Russia

Protein-directed fluorescent Au nanoclusters have been widely studied owing to their potential applications in sensing,imaging,and drug and gene ***,the use of nanoclusters in drug delivery is limited by low cellular *** this study,human serum albumin-directed Au nanoclusters served as building blocks to obtain protein nanoparticles by *** nanoparticles had a decent quantum yield(QY),high colloidal stability and low cytotoxicity,and they could be readily conjugated with biological *** cellular uptake of the Au nanoclusters and nanocluster-loaded protein nanoparticles were studied by confocal fluorescence *** of the protein-directed Au nanoclusters into 50–150-nm nanoparticles dramatically increased the cellular uptake.

关键词： Au nanoclusters albumin nanoparticles uptake fluorescence

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Polyphosphatase PPN1 of Saccharomyces cerevisiae Is a Deoxyadenosine Triphosphate Phosphohydrolase

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Advances in Enzyme Research 2016年第4期4卷 144-151页

作者： Nadezhda Andreeva Ludmila Trilisenko Mikhail Eldarov Tatiana Kulakovskaya skryabin institute of biochemistry and physiology of microorganisms Russian Academy of Sciences Pushchino Russia institute of Bioengineering Centre of Biotechnology Russian Academy of Sciences Moscow Russia

The Saccharomyces cerevisiae polyphosphatase PPN1 (uniprot/Q04119) degrades inorganic polyphosphates both by cleaving Pi from the chain end and by fragmenting long-chain polymers into shorter ones. In this study, we have found a new activity of this protein: it releases phosphate from dATP. The dATP phosphohydrolase activity of pure PPN1 was ~7-fold lower compared to the exopolyphosphatase activity. This activity was strongly stimulated by Co2+ ions, as well as by ammonium ions, and inhibited by heparin and pyrophosphate similar to the exopolyphosphatase activity of PPN1. The Km value for dATP was 0.88 ± 0.14 mM. The dATP phosphohydrolase activity in the cells of PPN1-overexpressing yeast strain was several-fold higher than that in the parent strain. The other exopolyphosphatase of S. cerevisiae, PPX1, did not split Pi from dATP.

关键词： Polyphosphate Yeast PPN1 Polyphosphatase Deoxyadenosine Triphosphate Phos-phohydrolase dATP

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Characteristics of Killer Toxin of the Yeast <i>Cryptococcus pinus</i>

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Journal of Biosciences and Medicines 2019年第4期7卷 73-82页

作者： Ekaterina Kulakovskaya Anton Zvonarev Vasilina Farofonova skryabin institute of biochemistry and physiology of microorganisms Russian Academy of Sciences Federal Research Center “Pushchino Research Center for Biological Studies” Russian Academy of Sciences Pushchino Russia

This work was aimed to study of fungicidal activity and mechanism of action of the killer toxin of Cryptococcus pinus VKM Y-2958. A killer-toxin preparation was obtained and partly purified from the culture liquid of the yeast Cryptococcus pinus. Its fungicidal activity against the pathogenic yeast Filobasidiella neoformans was assessed by assay of target cells growth inhibition and propidium iodide staining. The preparation contained three peptides with molecular masses of about 11 - 13 kDa, which did not show similarities in the mass spectrometric analysis. When frozen, the killer-toxin preparation remained active for several years. The killer toxin of Cr. pinus exhibits the high fungicidal activity against the known pathogen F. neoformans, is stable during long-term storage, and its mechanism of action is based on damaging target cell membranes. The killer toxin was obtained by a simple procedure and is of interest for the development of novel fungicidal preparations.

关键词： Cryptococcus pinus Filobasidiella neoformans Killer Toxin Fungicide Membrane-Damaging Activity

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