Biosensors Using Free and Immobilized Cells of Luminous Bacteria

The technologies of receiving free and immobilized photobacteria cells for biomonitoring of toxins are considered. The mechanisms of interaction of toxins with photobacteria are observed. The main attention is paid to the immobilized procedures and structures of carriers. Data on poly(vinyl)alcohol (PVA) cryogel immobilization of different strains of photobacteria are presented. It is established that intensity and stability of light emission of PVA cells is competently controlled by: (1) intensity and persistence of a luminescent cycle using bacterial strain; (2) type of the carrier and the composition of the gel-formation medium; (3) freeze-thawing procedures; and (4) physical and chemical conditions of storage and application. The developed technology of cryogenic gel formation has kept the survival of luminous bacteria in the carrier practically at 100% without the introduction of additional cryoprotecting agents and procedures of a light induction. With storage at −80°C, bioluminescent activity remained without changes about 2 years. Using the immobilized preparations of biosensor, the discrete and continuous analysis of heavy metals, chlorophenols, and pesticides is carried out. The sensitivity of free and immobilized cells to the chosen toxicants is approximately identical. The continuous monitoring of toxicant conditions is optimized

Višestruke mogućnosti primjene novih biokatalizatora pri enzimskoj obradi otpadnih voda prehrambene industrije

The cells of filamentous fungus R. oryzae entrapped in the polyvinyl alcohol cryogelare capable of producing various extracellular hydrolytic enzymes (proteases, amylases, lipases) and are used for the treatment of complex wastewaters of food industry. Five types of media simulating the wastewater of various food enterprises were treated under batch conditions for 600 h. Fats containing mostly residues of unsaturated fatty acids, as well as casein, glucose, sucrose, starch, soybean flour and various salts were the main components of the treated wastewaters. The immobilized cells concurrently possessed lipolytic, amylolytic and proteolytic activities. The level of each enzymatic activity depended on the wastewater content. The physiological state of immobilized cells was monitored by bioluminescent method. The intracellular adenosine triphosphate (ATP) concentration determined in the granules with immobilized cells was high enough and almost constant for all the period of biocatalyst application confirming thereby the active metabolic state of the cells. The study of mechanical strength of biocatalyst granules allowed revealing the differences in the values of modulus of biocatalyst elasticity at the beginning and at the end of its use for the wastewater treatment. The decrease in chemical oxygen demand of the tested media after their processing by immobilized biocatalyst was 68–79 % for one working cycle.Stanice filamentozne gljive Rhizopus oryzae ugrađene u kriogel polivinilnog alkohola mogu proizvesti razne ekstracelularne hidrolitičke enzime (proteaze, amilaze, lipaze), pa se koriste u obradi otpadnih voda prehrambene industrije. Pet vrsta podloga, koje simuliraju otpadne vode raznih prehrambenih poduzeća, obrađeno je u uvjetima šaržnog uzgoja tijekom 600 sati. Glavni su sastojci obrađenih otpadnih voda masti (najviše ostaci nezasićenih masnih kiselina), kazein, glukoza, saharoza, škrob, sojino brašno i razne soli. Utvrđeno je da imobilizirane stanice istodobno imaju lipolitička, amilolitička i proteolitička svojstva. Fiziološko stanje imobiliziranih stanica praćeno je pomoću bioluminiscentne metode. Intracelularna koncentracija adenozin trifosfata (ATP-a) u granulama s imobiliziranim stanicama je velika i skoro konstantna cijelo vrijeme primjene biokatalizatora, što dokazuje aktivni metabolički status stanica. Ispitivanje mehaničke čvrstoće granula biokatalizatora otkrilo je razliku u modulu elastičnosti prije i nakon obrade otpadnih voda. Smanjenje kemijske potrošnje kisika ispitanih podloga u jednom radnom ciklusu, nakon njihove obrade imobiliziranim biokatalizatorom, iznosilo je 68-79 %

Advanced Situation with Recombinant Toxins: Diversity, Production and Application Purposes

Today, the production and use of various samples of recombinant protein/polypeptide toxins is known and is actively developing. This review presents state-of-the-art in research and development of such toxins and their mechanisms of action and useful properties that have allowed them to be implemented into practice to treat various medical conditions (including oncology and chronic inflammation applications) and diseases, as well as to identify novel compounds and to detoxify them by diverse approaches (including enzyme antidotes). Special attention is given to the problems and possibilities of the toxicity control of the obtained recombinant proteins. The recombinant prions are discussed in the frame of their possible detoxification by enzymes. The review discusses the feasibility of obtaining recombinant variants of toxins in the form of protein molecules modified with fluorescent proteins, affine sequences and genetic mutations, allowing us to investigate the mechanisms of toxins’ bindings to their natural receptors

Enzymes for Detoxification of Various Mycotoxins: Origins and Mechanisms of Catalytic Action

Mycotoxins are highly dangerous natural compounds produced by various fungi. Enzymatic transformation seems to be the most promising method for detoxification of mycotoxins. This review summarizes current information on enzymes of different classes to convert various mycotoxins. An in-depth analysis of 11 key enzyme mechanisms towards dozens of major mycotoxins was realized. Additionally, molecular docking of mycotoxins to enzymes’ active centers was carried out to clarify some of these catalytic mechanisms. Analyzing protein homologues from various organisms (plants, animals, fungi, and bacteria), the prevalence and availability of natural sources of active biocatalysts with a high practical potential is discussed. The importance of multifunctional enzyme combinations for detoxification of mycotoxins is posed

Enzymes, Reacting with Organophosphorus Compounds as Detoxifiers: Diversity and Functions

Organophosphorus compounds (OPCs) are able to interact with various biological targets in living organisms, including enzymes. The binding of OPCs to enzymes does not always lead to negative consequences for the body itself, since there are a lot of natural biocatalysts that can catalyze the chemical transformations of the OPCs via hydrolysis or oxidation/reduction and thereby provide their detoxification. Some of these enzymes, their structural differences and identity, mechanisms, and specificity of catalytic action are discussed in this work, including results of computational modeling. Phylogenetic analysis of these diverse enzymes was specially realized for this review to emphasize a great area for future development(s) and applications

“Deceived” Concentrated Immobilized Cells as Biocatalyst for Intensive Bacterial Cellulose Production from Various Sources

A new biocatalyst in the form of Komagataeibacter xylinum B-12429 cells immobilized in poly(vinyl alcohol) cryogel for production of bacterial cellulose was demonstrated. Normally, the increased bacteria concentration causes an enlarged bacterial cellulose synthesis while cells push the polysaccharide out to pack themselves into this polymer and go into a stasis. Immobilization of cells into the poly(vinyl alcohol) cryogel allowed “deceiving” them: bacteria producing cellulose pushed it out, which further passed through the pores of cryogel matrix and was accumulated in the medium while not covering the cells; hence, the latter were deprived of a possible transition to inactivity and worked on the synthesis of bacterial cellulose even more actively. The repeated use of immobilized cells retaining 100% of their metabolic activity for at least 10 working cycles (60 days) was performed. The immobilized cells produce bacterial cellulose with crystallinity and porosity similar to polysaccharide of free cells, but having improved stiffness and tensile strength. Various media containing sugars and glycerol, based on hydrolysates of renewable biomass sources (aspen, Jerusalem artichoke, rice straw, microalgae) were successfully applied for bacterial cellulose production by immobilized cells, and the level of polysaccharide accumulation was 1.3–1.8-times greater than suspended cells could produce

Optimization of the Use of His6-OPH-Based Enzymatic Biocatalysts for the Destruction of Chlorpyrifos in Soil

Applying enzymatic biocatalysts based on hexahistidine-containing organophosphorus hydrolase (His6-OPH) is suggested for the decomposition of chlorpyrifos, which is actively used in agriculture in many countries. The application conditions were optimized and the following techniques was suggested to ensure the highest efficiency of the enzyme: first, the soil is alkalinized with hydrated calcitic lime Ca(OH)2, then the enzyme is introduced into the soil at a concentration of 1000 U/kg soil. Non-equilibrium low temperature plasma (NELTP)-modified zeolite is used for immobilization of the relatively inexpensive polyelectrolyte complexes containing the enzyme His6-OPH and a polyanionic polymer: poly-l-glutamic acid (PLE50) or poly-l-aspartic acid (PLD50). The soil’s humidity is then increased up to 60–80%, the top layer (10–30 cm) of soil is thoroughly stirred, and then exposed for 48–72 h. The suggested approach ensures 100% destruction of the pesticide within 72 h in soils containing as much as 100 mg/kg of chlorpyrifos. It was concluded that using this type of His6-OPH-based enzyme chemical can be the best approach for soils with relatively low humus concentrations, such as sandy and loam-sandy chestnut soils, as well as types of soil with increased alkalinity (pH 8.0–8.4). Such soils are often encountered in desert, desert-steppe, foothills, and subtropical regions where chlorpyrifos is actively used