Constitutive chitosanase from Bacillus thuringiensis B-387 and its potential for preparation of antimicrobial chitooligomers

• Published in: World journal of microbiology & biotechnology 2022-10, Vol.38 (10), p.167-167, Article 167
• Main Authors: Aktuganov, Gleb E., Safina, Violetta R., Galimzianova, Nailya F., Gilvanova, Elena A., Kuzmina, Lyudmila Yu, Melentiev, Alexander I., Baymiev, Andrei H., Lopatin, Sergey A.
• Format: Article
• Language: English
• Subjects: Antibacterial activity; Antimicrobial agents; Applied Microbiology; Biochemistry; Biomedical and Life Sciences; Biotechnology; Cation exchanging; Cation-exchange chromatography; Cellulose; Chitin; Chitosan; Chitosanase; Colloid chitin; Deacetylation; Environmental Engineering/Biotechnology; Enzymes; Enzymolysis; Fungicides; Gel electrophoresis; Glucans; Life Sciences; Microbiology; Molecular weight; Original Paper; pH effects; Polysaccharides; Reaction products; Saccharides; Sodium lauryl sulfate; Substrates; Ultrafiltration
• ISSN: 0959-3993; 1573-0972
• DOI: 10.1007/s11274-022-03359-5

The article proves the ability of the entomopathogenic strain B. thuringiensis var. dendrolimus B-387 to high the constitutive production (3–12.5 U/mL) of extracellular chitosanase, that was found for the first time. The enzyme was purified in 94-fold by ultrafiltration, affinity sorption and cation-exchange chromatography and characterized biochemically. The molecular mass of the chitosanase determined using SDS-PAGE is 40 kDa. Temperature and pH-optima of the enzyme are 55  ° C and pH 6.5, respectively; the chitosanase was stable under 50–60  ° C and pH 4–10.5. Purified chitosanase most rapidly (V max  ~ 43 µM/mL × min, K M  ~ 0.22 mg/mL, k cat  ~ 4.79 × 10 4  s −1 ) hydrolyzed soluble chitosan of the deacetylation degree (DD) 85% by endo-mode, and did not degrade colloidal chitin, CM-cellulose and some other glucans. The main reaction products of the chitosan enzymolysis included chitobiose, chitotriose and chitotetraose. In addition to small chitooligosaccharides (CHOs), the studied chitosanase also generated low-molecular weight chitosan (LMWC) with average M w in range 14–46 kDa and recovery 14–35%, depending on the enzyme/substrate ratio and incubation temperature. In some cases, the chitosan (DD 85 and 50%) oligomers prepared using crude chitosanase from B . thuringiensis B-387 indicated higher antifungal and antibacterial activities in vitro in comparison with the initial polysaccharides. The data obtained indicate the good prospect of chitosanase B-387 for the production of bioactive CHOs. Graphical abstract