Increasing chitosanase production in Bacillus cereus by a novel mutagenesis and screen method

Abstract

Chitosan hydrolysis by chitosanase is one of the most effective methods to produce chitosan oligosaccharides. One of the prerequisites of enzyme fermentation production is to select and breed enzyme-producing cells with good performance. So in the process of fermentation production, the low yield of chitosanase cannot meet the current requirement. In this paper, a strain producing chitosanase was screened and identified, and a novel mutagenesis system (Atmospheric and Room Temperature Plasma (ARTP)) was selected to increase the yield of chitosanase. Then, the fermentation medium was optimized to further improve the enzyme activity of the strain. A strain of Bacillus cereus capable of producing chitosanase was screened and identified from soil samples. A mutant strain of B.cereus was obtained by Atmospheric and Room Temperature Plasma mutagenesis and bioscreening method, and chitosanase activity was 2.49 folds that of the original bacterium. After an optimized fermentation medium, the enzyme activity of the mutant strain was 1.47 folds that of the original bacterium. Combined with all the above optimization experiments, the enzyme activity of mutant strain increased by 3.66 times. The results showed that the Atmospheric and Room Temperature Plasma mutagenesis and bioscreening method could significantly increase the yield of chitosanase in B.cereus, and had little effect on the properties of the enzyme. These findings have potential applications in the mutagenesis of other enzyme-producing microorganisms.

Fungicides alter the distribution and diversity of bacterial and fungal communities in ginseng fields

“…Similarly, Huang et al [ 22 ] observed significantly increased in Arthrobacter abundance in dazomet-treated soil, which might be attributed by Arthrobacter ability to degrade dazomet as a nutrient source. And Bacillus is favorable to rhizobacteria that absorbs nutrients and promotes plant yield, and also potentially contributed as antimicrobial agents owing to the production of a variety of antibiotics, including bacitracin and lipopeptides, polyketones, and lipopeptides [ 23 , 24 ], further used in organic biological products to combat plant pathogens and inhibit various soil-borne pathogens.

Figure 1.

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“…Thus, the obvious degenerate tendency of Proteobacteria abundance after apply fungicides in this study revealed the positive effect of fungicide on beneficial microbial. Conversely, Acidobacteria was able to oxidize iron and deposit and caused surface rusty in ginseng root [21], the relative abundance (RA) of Acidobacteria was reduced from 27.76 to 7.14% and minimum in E. However, the phylum Actinobacteria was contributed as decomposers of organic matter and enemies of soil-borne pathogens or act as biocontrol agents to control plant diseases transmitted by soil and seeds [6,22] And Bacillus is favorable to rhizobacteria on absorb nutrients and promote plant yield, also potentially contributed as antimicrobial agents owing to produced variety of antibiotics, including bacitracin and lipopeptides, polyketones and lipopeptides [24,25], further used in organic biological products to combat plant pathogens and inhibit various soil-borne pathogens.…”

High-Throughput Screening Techniques for the Selection of Thermostable Enzymes

“…19 Currently, rational design is favored as the primary method for enzyme improvement; however, directed evolution remains preferable when dealing with proteins with unknown structures or functions. 20 Rapid screening technology is essential for all these approaches in order to identify enzymes with desired properties efficiently, particularly in the case of directed evolution, which generates vast variant libraries.…”

“…It employs various combinations of methods to improve efficiency by reducing library size while preserving necessary diversity provided by directed evolution. , Combinatorial active-site saturation test (CAST) and iterative saturation mutagenesis (ISM) have emerged as prominent approaches in semirational design for enhancing enzyme activity and selectivity . Currently, rational design is favored as the primary method for enzyme improvement; however, directed evolution remains preferable when dealing with proteins with unknown structures or functions . Rapid screening technology is essential for all these approaches in order to identify enzymes with desired properties efficiently, particularly in the case of directed evolution, which generates vast variant libraries.…”

Covalent immobilization of Dothideomycetes sp. NRC-SSW chitosanase and its application in chitosan hydrolysis

“…Chemical hydrolysis was the commercial applied method for chitosan de-polymerization but in the last few years the use of enzymes has been suggested as an alternative method from the perspective of employed mild conditions, environmental protection and high yield of the resulted product. The cost, availability and specificity of the applied enzyme are the main variables that have to be considered for the industrial applications of chitosan enzymatic hydrolysis [10][11][12][13]. Chitosanases are the specific chitosanolysis enzymes.…”