Genetic Improvement of Bacillus licheniformis Strains for Efficient Deproteinization of Shrimp Shells and Production of High-Molecular-Mass Chitin and Chitosan

• Published in: Applied and Environmental Microbiology Vol. 76; no. 24; pp. 8211 - 8221
• Main Authors: Hoffmann, Kerstin, Daum, Gabriele, Köster, Marina, Kulicke, Werner-Michael, Meyer-Rammes, Heike, Bisping, Bernward, Meinhardt, Friedhelm
• Format: Journal Article
• Language: English
• Published: Washington, DC American Society for Microbiology 01.12.2010; American Society for Microbiology (ASM)
• Subjects: Animals; Bacillus - genetics; Bacillus - metabolism; Bacillus licheniformis; Bacteria; Biochemistry; Biological and medical sciences; Biotechnology; Carbohydrates; Chitin; Chitin - chemistry; Chitin - isolation & purification; Chitin - metabolism; chitinase; chitosan; Chitosan - chemistry; Chitosan - isolation & purification; Chitosan - metabolism; Chromatography, Gel; Fermentation; Fundamental and applied biological sciences. Psychology; gel chromatography; Genes, Bacterial; genetic improvement; Genetics; Genotype & phenotype; lipids; Lipids - analysis; Microbiology; Molecular Weight; NMR; Nuclear magnetic resonance; nuclear magnetic resonance spectroscopy; operon; Organisms, Genetically Modified - genetics; Organisms, Genetically Modified - metabolism; Penaeidae - microbiology; Polyglutamic Acid - metabolism; proteolysis; Saccharides; Sequence Deletion; Shellfish; shrimp; Spectrum analysis; viscometry; viscosity; Bacillus licheniformis; Chitin; Bacillaceae; Deproteinization; Macrura; Crustacea; Bacillales; Arthropoda; Genetic improvement; Production; Bacteria; Decapoda; Chitosan; Invertebrata; Molecular mass; Shrimp
• ISSN: 0099-2240; 1098-5336; 1098-5336; 1098-6596
• DOI: 10.1128/AEM.01404-10

By targeted deletion of the polyglutamate operon (pga) in Bacillus licheniformis F11, a derivative form, F11.1 (Δpga), was obtained that, along with lacking polyglutamate (PGA) formation, displayed enhanced proteolytic activities. The phenotypic properties were maintained in a strain in which the chiBA operon was additionally deleted: F11.4 (ΔchiBA Δpga). These genetically modified strains, carrying the Δpga deletion either alone (F11.1) or together with the ΔchiBA (F11.4) deletion, were used in fermentations (20-liter scale) aiming at the deproteinization of shrimp shells in order to obtain long-chain chitin. After chemical deacetylation, the resulting chitosan samples were analyzed by nuclear magnetic resonance spectroscopy, size exclusion chromatography, and viscometry and compared to a chitosan preparation that was produced in parallel by chemical methods by a commercial chitosan supplier (GSRmbH). Though faint lipid impurities were present in the fermented polysaccharides, the viscosity of the material produced with the double-deletion mutant F11.4 (Δpga ΔchiBA) was higher than that of the chemically produced and commercially available samples (Cognis GmbH). Thus, enhanced proteolytic activities and a lack of chitinase activity render the double mutant F11.4 a powerful tool for the production of long-chain chitosan.