RESUMO
The ability to produce extracellular chitosanase (EC 3.2.1.132) was found by plate assays in 18 (23%) out of 77 crystalliferous strains of Bacillus thuringiensis. The best chitosanase producer was selected after the growth chosen in a liquid medium with colloidal chitosan as carbon source. Enzyme production was optimized (a 4-d incubation at 32 degrees C with shaking in a medium of pH 6.5 with 4% colloidal chitosan) and the enzyme was partially characterized. This is the first report on the chitosanase of B. thuringiensis.
Assuntos
Bacillus thuringiensis/enzimologia , Quitina/análogos & derivados , Glicosídeo Hidrolases/metabolismo , Quitina/metabolismo , Quitosana , Cromatografia em Gel , Eletroforese em Gel de Poliacrilamida , Inibidores Enzimáticos/farmacologia , Estabilidade Enzimática , Glicosídeo Hidrolases/isolamento & purificação , Concentração de Íons de Hidrogênio , Peso Molecular , Controle Biológico de Vetores , TemperaturaRESUMO
A simple and sensitive method based on the use of colloidal chitin stained with Remazol Brilliant Blue R (RBB) is proposed to evaluate chitinase activity. If this colloidal-stained substrate is included as a carbon source in a liquid medium, this technique allows the selection or the comparison of chitinolytic microorganisms. The colloidal substrate is proportionally solubilized and the dye released is spectrophotometrically quantified at 595 nm. The procedures used for the staining and fixing of RBB in the colloidal chitin, and a comparison with the commercial substrate chitin-azure, are presented. The influence of several physicochemical and enzymatic parameters on the release of dyes is also shown. Both stained substrates were used for studying the effect of pH, substrate concentration, temperature and time on the chitinase reaction of Bacillus thuringiensis Bt-107.
