Production Chitosanase Strain, Fermentation Conditions Optimization And Enzymatic Properties

A strain with chitosanase activity is isolated from soil samples. After being mutated by uv for 6minutes and 1.00mg/mL concentration of NTG treatments for 30 minutes, it is named as H1. The optimal compositions of the medium for H1 strain producing chitosanase are as follows(%, w/v): (NH₄)₂SO₄1.00, yeast extract 0.30, peptone 0.50, K₂HPO₄0.070, KH₂PO₄0.030, NaCl 0.50, MgSO₄ .7H₂O 0.050, glucose 0.10, powder chitosan 1.00, the initial pH5.00. The optimal cultivation conditions for H1 producing chitosanase are3.0% (v/v) amount of inoculation , 30℃ incubating temperature and shaking cultivation at 150rpm for 96 hours. The chitosanase activity of fermented broth could reach 9U/mL when H1 is incubated in flask under the optimal cultivition conditions. The chitosanase from H1 strain is an inducible and secreting extracellar enzyme. The chitosanase of the strain is extracted by ammonium sulfate precipitation (30-70% saturation) , and purifies by ion-exchange chromatography (Q-Sepharose Fast Flow colum) and gel filtration (Sephadex G-75 colum), SDS-PAGE shows a single band for the purified chitosanase and the molecular weight is estimated to be 43kD. The optimal temperature and pH for hydrolysis of the chitosanase are 60℃ and 5.2, respectively. Mn²+ , Fe²+and Mg²+enhance the enzyme activity significantly, whereas Cu²+ ,Zn²+ inhibit the enzyme activity. The enzyme shows activity for hydrolysis of chitosan, chitin and carboxymethyl cellulose.