卷 55, 编号 8 (2019)

This work describes the properties of a new protein, a modification of GroEL apical domain designed to be a leader in fusion systems. This polypeptide leader demonstrates a high level of expression in a bacterial system; it is soluble and retains its solubility during standard biochemical manipulations. The secondary structure of the protein and its thermostability, as well as the protein solubility, were studied in a wide temperature range. To simplify the subsequent purification of the target protein, the possibility of its chemical cleavage from the fused protein by methionine residues with cyanogen bromide is provided.

The expression of four genes of bacterial endo-β-l,3(4)-D-glucanases from B. pumilus Bg57 (VKPM В-13195), Paenibacillus polymyxa Bg23 (VKPM В-4056), Bacillus subtilis Bg11 (VKPM B-13178), and Bacillus amyloliquefaciens Bg76 (VKPM B-13184), which are members of the family of GH16 glycohydrolases, in the expression system of Pichia pastoris are compared. Technologically valuable characteristics of the recombinant β-glucanases, such as the specific activity, pH and thermal stability, temperature and pH optima, resistance to digestive enzymes, and substrate specificity, were studied. It was shown that the enzymes from B. pumilus and P. polymyxa have the highest biotechnological potential for the creation of β‑glucanase producers based on recombinant P. pastoris yeast strains for further use in fodder production.

The heterologous expression, isolation, and characterization of a novel xylanase from Paenibacillus brasilensis are described. The xyl1 gene from the Paenibacillus brasilensis strain X1 VKPM B-13092, which consists of 639 nucleotides, encodes a secreted endo-1,4-β-xylanase (EC 3.2.1.8) containing 184 amino acids and 28 residues of the putative signal peptide in the N-terminal region. The nucleotide sequence of the xyl1 gene and the amino acid sequence of the mature Xyll protein have the greatest homology with the Bacillus subtilis endo-1,4-β-xylanase sequences (78 and 83%, respectively). A gene fragment encoding the mature protein was expressed in Pichia pastoris. The purified recombinant Xyl1 enzyme was able to use birch xylan and arabinoxylan as substrates. With birch xylan, the optimal pH for the enzymatic reaction was 6.0, the optimal temperature was 40–50°C, and K_m and V_max, were equal to 1.1288 mg/mL and 5124.3 μmol/(min mg), respectively. The recombinant Xyl1 protein showed high pH and thermal stability, and the resistance to digestive enzymes and xylanase protein inhibitors from cereals. It was also shown that Mn²⁺ and Со²⁺ ions stimulate enzyme activity.

The protection of plants from diseases caused by various pathogens is an economically and socially important problem; the losses in crop production constitute 20% of the harvest in different parts of the world. The use of chemical pesticides is the main method of plant protection. However, chemical preparations have a number of serious disadvantages. Biologicals for plant protection (BPP) are currently being developed more intensely. The world biggest chemical companies, BASF, Bayer and Syngenta, show great interest in the market for preparations of biological control. The data on markets of biologicals for the protection of agricultural plants in various parts of the world (North America, Europe, China, Latin America and Russia) are presented. According to expert data, the biological markets in these areas (with the exception of Russia) will exceed $1 billion by 2025. In Russia, only 0.3% of agricultural land is treated with biological products. The review contains data on the use of biofungicides obtained at GosNIIgenetika for the protection of wheat, potato, and vegetable crops. A high efficiency of the biofungicides has been demonstrated.