Papers by Valentina Kratasyuk
Bioinformatics (Oxford, England), Jan 26, 2016
Bacterial luciferases are heterodimeric enzymes that catalyze a chemical reaction, so called biol... more Bacterial luciferases are heterodimeric enzymes that catalyze a chemical reaction, so called bioluminescence, which causes light emission in bacteria. Bioluminescence is vastly used as a reporter system in research tools and commercial developments. However, the details of the mechanisms that stabilize and transform the reaction intermediates as well as differences in the enzymatic kinetics amongst different bacterial luciferases remain to be elucidated. Amino acid sequences alignments for 21 bacterial luciferases (both α- and β-subunits) were analyzed. For α-subunit, containing the enzyme active center, 48 polymorphic amino acid positions were identified. According to them, the sequences fell into two distinct groups known as slow and fast based on the decay rate of the bioluminescence reaction. The differences in the enzyme active site induced by structural polymorphism are analyzed. [email protected] SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics o...
Izvestiya of Altai State University, 2013
Sensors and Actuators B: Chemical, 2015
The biomodule of bioluminescent biosensor based on a coupled enzyme system NADH:FMNoxidoreductase... more The biomodule of bioluminescent biosensor based on a coupled enzyme system NADH:FMNoxidoreductase and luciferase, co-immobilized with substrates in dried starch or gelatin gels, has been developed. We studied the impact of several stabilizers-dithiothreitol (DTT), bovine serum albumin (BSA) and mercaptoethanol (ME) on the biomodule's activity, storage stability and sensitivity to toxic substances. The inclusion of stabilizers increases the activity of the biological module by more than 150%. To achieve the combination of high activity, prolonged storage time and acute sensitivity to toxic substances within maximum permissible concentration we used starch gel as a carrier adding 100 M DTT to the immobilized preparation. The gelatin-based biological module had greater storage stability than the starch-based one but demonstrated less sensitivity to toxic substances. . Prof. Kratasyuk pioneered enzymatic bioluminescent biotests and coimmobilization techniques for luminescent enzyme systems. Her research focuses on bioluminescent analysis and ecological biophysics with a particular interest in design of bioluminescent enzymatic assays for monitoring the wide range of biological and ecological systems. She is the author of more than one hundred papers and fifteen patents on bioluminescence and biosensors.
Journal of biomolecular structure & dynamics, 2015
Bioluminescence studies have been incorporated into the six-week Kennedy Space Center Spaceflight... more Bioluminescence studies have been incorporated into the six-week Kennedy Space Center Spaceflight and Life Sciences Training Program (SLSTP). The SLSTP program is designed to attract the "best and brightest" US and Canadian students to careers in space science and engineering. This six-week program was sponsored by NASA and implemented at the Kennedy Space Center (KSC). Dynamac and Bionetics Corporations provided the research projects that were integrated in a program designed and assesed by a consortium of post-secondary institutions.
Bioluminescence and Chemiluminescence - Chemistry, Biology and Applications - Proceedings of the 14th International Symposium, 2007
Bulletin of experimental biology and medicine, 2003
The integral bioluminescent biotest with lyophilized fluorescent bacteria was used for monitoring... more The integral bioluminescent biotest with lyophilized fluorescent bacteria was used for monitoring of LPO processes in tissue extracts and serum of rats exposed to stress. A relationship between the content of MDA (LPO indicator) and fluorescence of bacteria was observed in all biological samples.
Bulletin of experimental biology and medicine, 2003
We studied the possibility of evaluation of the intensity of pathological oxidative processes in ... more We studied the possibility of evaluation of the intensity of pathological oxidative processes in rat liver using an integral bioluminescent test with controlled perfusion of the isolated organ. The test revealed a significant correlation between the level of TBA-reactive products and bioluminescence intensity.
International Journal of Salt Lake Research, 1999
Field Analytical Chemistry & Technology, 1998
Analytical and Bioanalytical Chemistry, 2014
We have studied the effects of a gel-like environment on the characteristics of enzyme preparatio... more We have studied the effects of a gel-like environment on the characteristics of enzyme preparations based on the coupled enzyme system of luminous bacteria, NADH:FMN-oxidoreductase-luciferase, to design a stable immobilizing reagent for bioluminescent analysis. Natural polymers, gelatin and starch, were used to create a viscous, structured microenvironment. The stability of the coupled enzyme system to such physical and chemical environmental factors as temperature, pH, and ionic strength in gelatin and starch-containing media was examined. It was shown that both gelatin and starch have a stabilizing effect on the enzymes of luminous bacteria under specific conditions. In particular, the enzymes' activity is increased twofold in the presence of 1 and 5 % of gelatin at 20°C and 25°C, respectively (temperatures lower than the gel point). Also, the acceptable pH range of the coupled enzyme system expands into the alkaline region and becomes 6.8-8.1. Stabilization at low ionic strength (0.01-0.06 mol L −1 ) is observed. At the same time, microenvironments based on either gelatin or starch do not change the enzymes' thermal inactivation rate constants in the temperature range from 25 to 43°C. Finally, gelatin and starch are suitable for development of a reagent for immobilization of enzymes which would be stable and resistant to physical and chemical environmental conditions.
Advances in Biochemical Engineering/Biotechnology, 2014
This review examines the general principles of bioluminescent enzymatic toxicity bioassays and de... more This review examines the general principles of bioluminescent enzymatic toxicity bioassays and describes the applications of these methods and the implementation in commercial biosensors. Bioluminescent enzyme system technology (BEST) has been proposed in the bacterial coupled enzyme system, wherein NADH:FMN-oxidoreductase-luciferase substitutes for living organisms. BEST was introduced to facilitate and accelerate the development of cost-competitive enzymatic systems for use in biosensors for medical, environmental, and industrial applications. For widespread use of BEST, the multicomponent reagent ''Enzymolum'' has been developed, which contains the bacterial luciferase, NADH:FMNoxidoreductase, and their substrates, co-immobilized in starch or gelatin gel. Enzymolum is the central part of Portable Laboratory for Toxicity Detection (PLTD), which consists of a biodetector module, a sampling module, a sample preparation module, and a reagent module. PLTD instantly signals chemicalbiological hazards and allows us to detect a wide range of toxic substances. Enzymolum can be integrated as a biological module into the portable biodetectorbiosensor originally constructed for personal use. Based on the example of Enzymolum and the algorithm for creating new enzyme biotests with tailored characteristics, a new approach was demonstrated in biotechnological design and construction. The examples of biotechnological design of various bioluminescent methods for ecological monitoring were provided. Possible applications of enzyme bioassays are seen in the examples for medical diagnostics, assessment of the effect of physical load on sportsmen, analysis of food additives, and in practical courses for higher educational institutions and schools. The advantages of enzymatic assays are their rapidity (the period of time required does not exceed 3-5 min), high E. Esimbekova Á V. Kratasyuk (, Ó Springer-Verlag Berlin Heidelberg 2014 67 sensitivity, simplicity and safety of procedure, and possibility of automation of ecological monitoring; the required luminometer is easily available.
Photochemical & Photobiological Sciences, 2007
A review of the mechanisms of the exogenous redox compounds influence on the bacterial coupled en... more A review of the mechanisms of the exogenous redox compounds influence on the bacterial coupled enzyme system: NAD(P)H:FMN-oxidoreductase-luciferase has been done. A series of quinones has been used as model organic oxidants. The three mechanisms of the quinones' effects on bioluminescence were suggested: (1) inhibition of the NADH-dependent redox reactions; (2) interactions between the compounds and the enzymes of the coupled enzyme system; and (3) intermolecular energy migration. The correlation between the kinetic parameters of bioluminescence and the standard redox potential of the quinones proved that the inhibition of redox reactions was the key mechanism by which the quinones decrease the light emission intensity. The changes in the fluorescence anisotropy decay of the endogenous flavin of the enzyme preparations showed the direct interaction between quinones and enzymes. It has been demonstrated that the intermolecular energy migration mechanism played a minor role in the effect of quinones on the bioluminescence. A comparative analysis of the effect of quinones, phenols and inorganic redox compounds on bioluminescent coupled enzyme systems has been carried out.
Luminescence, 2007
Prototype technologies of a bioluminescent signal system (BSS) based on the luminous bacterium Ph... more Prototype technologies of a bioluminescent signal system (BSS) based on the luminous bacterium Photobacterium phosphoreum and three enzymatic bioluminescence systems have been proposed for detecting and signalling the presence of toxicants in water systems. A number of pesticides, mostly known as poisonous substances, similar in their structures and physicochemical properties, have been taken as model compounds of chemical agents. The effect of toxicants (organophosphates, derivatives of dithiocarbamide acid, and pyrethroid preparations) on the bioluminescence of the four systems has been analysed. EC 50 and EC 80 have been determined and compared to the maximum permissible concentration for each of the analysed substances. The triple-enzyme systems with ADH and trypsin have been shown to be more sensitive to organophosphorous compounds (0.13-11 mg/L), while the triple-enzyme system with trypsin is highly sensitive to lipotropic poison, a derivative of dithiocarbamine acid (0.03 mg/L). Sensitivities of the triple-enzyme systems to pyrethroid preparations are similar to those of luminous bacteria (0.9-5 mg/L). The results can be used to construct an alarm-test bioluminescence system for detecting chemical toxicants, based on intact bacteria or enzyme systems. Bioluminescent signal system for chemical toxicants in water ORIGINAL RESEARCH 209 E. Vetrova et al. Bioluminescent signal system for chemical toxicants in water ORIGINAL RESEARCH 211 E. Vetrova et al.
Luminescence, 1999
We describe our experience with laboratory courses in enzymology based on the phenomenon of biolu... more We describe our experience with laboratory courses in enzymology based on the phenomenon of bioluminescence. The soluble and immobilized enzymes of luminous bacteria are used and the practical enzymological course consists of four main courses: (1) training in measuring the activities of soluble and immobilized enzymes; (2) the investigation of kinetic characteristics (kinetic constants) and enzyme-substrate and enzyme-inhibitor interactions in the bacterial bioluminescent reaction; (3) The testing of physico-chemical characteristics of enzymes (pH, temperature, ion strength, etc.); (4) the effect of inhibitors on enzymes. Training is possible in groups of about ten persons. Our practice work has been introduced in the biological, pedagogical and physical departments of Krasnoyarsk State University. Students of the pedagogical department have created a popular and interesting series of laboratory works for high school children aged 14-17 years.
Luminescence, 2003
The study addressed the effects of redox-active compounds on trypsin activity. Series of organic ... more The study addressed the effects of redox-active compounds on trypsin activity. Series of organic oxidizers (quinones) and reducers (phenols) were chosen as model redox-active compounds. Trypsin activity was quantified by bioluminescent technique. Interactions of these compounds with trypsin were studied by fluorescent and light absorption methods. Luminescence intensity decay constants in the reduced nicotinamidadeninedinucleotide (NADH): flavinmononucleotide (FMN)-oxidoreductase (R)-luciferase (L)-trypsin (T) (R + L + T) triple-enzyme system were calculated and compared in the presence of different concentrations of quinones and phenols. The triple-enzyme system was shown to be sensitive to quinones and not sensitive to phenols. It has been found that the effects produced by quinones on the coupled enzyme system (R + L) and on the trypsin molecule (T) are not related. The conclusions were extrapolated to the properties of other proteases and antiproteases.
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Papers by Valentina Kratasyuk