Enzymatic Lysis of Living Microbial Cells: A Universal Approach to Calculating the Rate of Cell Lysis in Turbidimetric MeasurementsMatolygina, D.; Dushutina, N.; Ovchinnikova, E.; Eremeev, N.; Belogurova, N.; Atroshenko, D.; Smirnov, S.; Savin, S.; Tishkov, V.; Levashov, A.; Levashov, P.
2018 Moscow University Chemistry Bulletin
doi: 10.3103/S0027131418020104
The data of the turbidimetric measurement for the enzymatic lysis of various living bacterial cells are analyzed. A method for the correct recalculation of the turbidimetric data (–ΔA/Δt) into absolute values of the change in the concentration of living cells (–ΔCFU/Δt) is proposed. The dimensionless efficiency of cell lysis–(1/CFU0) · ΔCFU/Δt for various bacterial cells is calculated to correctly compare the efficiency of the action of different bacteriolytic factors on various bacterial cells.
A New Enzyme Preparation for Reducing the Viscosity of Whole-Grain Rye ExtractsRozhkova, A.; Miarzlou, D.; Bashirova, A.; Zorov, I.; Korotkova, O.; Shashkov, I.; Sinitsyn, A.
2018 Moscow University Chemistry Bulletin
doi: 10.3103/S0027131418020141
New recombinant strains based on the lower fungus Penicillium canescens are engineered to produce an extracellular enzyme complex that contains a homologous endo-1,4-β-xylanase E (XylE, EC 3.2.1.8). Expression of the gene encoding XylE is controlled by the promoter of the xylA gene, which encodes endo- 1,4-β-xylanase A. Comparative analysis of enzyme preparations (EPs) isolated from the culture liquid of the host strain (Penicillium canescens RN3-11-7) and the new recombinant P. canescens strains of the XylE series is performed. The specific activity of the EPs against a specific substrate (birch glucuronoxylan) is studied, and the qualitative and quantitative composition of the new EPs XylE-B5 and XylE-C1, as well as the RN3 EP from the host strain, is determined. The decrease in the reduced viscosity of native (not subjected to thermal inactivation) aqueous extracts of rye treated with XylE-B5 EP was double that in the case of treatment with RN3 EP, due to high content of inhibition-resistant XylE in the new EPs.
Enzymatic Hydrolysis of Cellulose Using Mixes of Mutant Forms of Cellulases from Penicillium verruculosumDotsenko, A.; Gusakov, A.; Rozhkova, A.; Volkov, P.; Korotkova, O.; Sinitsyn, A.
2018 Moscow University Chemistry Bulletin
doi: 10.3103/S0027131418020037
Cellulases are the major components of multienzyme systems applied in processes of bioconversion of renewable lignocellulosic feedstocks to various useful products. The hydrolytic efficiency of enzyme mixes based on recombinant wild-type endoglucanase II, cellobiohydrolases I and II from the Penicillium verruculosum fungus (in the presence of Aspergillus niger β-glucosidase) with mixes of mutant forms of these enzymes in the hydrolysis of cellulosic materials is compared, and the influence of temperature and substrate concentration on the glucose yield is studied. The mutant cellulases represented proteins, in which N-linked glycans were partially removed using site-directed mutagenesis. In the hydrolysis of microcrystalline cellulose and milled aspen wood by mixes of mutant cellulases, the yields of glucose after 24–72 h of an enzymatic reaction were higher by 31–38% and 11–27%, respectively, than those for the compositions based on the wild-type enzymes. The highest product concentrations, using mutant enzyme compositions, are achieved at 50°С when the hydrolysis temperature is varied in the range of 40 to 60°С. Increasing the substrate concentration in the reaction system from 5 to 50 g/L (while maintaining the enzyme dosage at the same level) led to a 2.6–2.8-fold increase in the glucose yield, accompanied by a decrease in the cellulose conversion degree.
Multifunctional Lipoamino Acid Derivatives with Potential Biological ActivityMarusova (Soloveva), V.; Zagitova, R.; Budanova, U.; Sebyakin, Yu.
2018 Moscow University Chemistry Bulletin
doi: 10.3103/S0027131418020098
This work aims to develop a number of multifunctional derivatives of lipoamino acids, which potentially have different biological activities. To achieve this aim, the hydrophilic-lipophilic balance for a number of cationic amphiphile structures based on L-glutamic acid and short diamines is theoretically calculated. The developed schemes and performed synthesis allowed us to produce preparative sample quantities for further physicochemical and biological tests. The performed calculations, measurements, and experiments on the liposomal dispersions of the synthesized compounds demonstrate the possibility of designing the same type of structures of an amphiphilic nature based on aliphatic amino acid derivatives and diamines, which can be used both in antibacterial therapy and as the delivery vehicles for genetic material.
Comparison of Thermal Stability of New Formate Dehydrogenases by Differential Scanning CalorimetryPometun, A.; Kleymenov, S.; Zarubina, S.; Kargov, I.; Parshin, P.; Sadykhov, E.; Savin, S.; Tishkov, V.
2018 Moscow University Chemistry Bulletin
doi: 10.3103/S002713141802013X
Formate dehydrogenases (FDHs) from different sources are systematically studied in our laboratory. Over the past few years, new genes of four FDHs from pathogenic bacterium Staphylococcus aureus (SauFDH), methylotrophic thermotolerant yeast Ogataea parapolymorpha (OpaFDH), yeast Saccharomyces cerevisiae (SceFDH), and moss Physcomitrella patens (PpaFDH) have been cloned and expressed in E. coli cells. By means of differential scanning calorimetry, a comparative study of thermal stability of new recombinant formate dehydrogenases and a number of FDHs from other sources has been performed. It was shown that two new enzymes, SauFDH and OpaFDH, are comparable to the FDH from Pseudomonas sp. 101 bacteria in their stability. SceFDH is the least stable FDH among the described formate dehydrogenases.