Purification and characterization of chitinase from Paenibacillus sp.

Abstract The chitinase‐producing bacteria Paenibacillus sp. was isolated from soil samples. The chitinase was purified successively by ammonia sulfate fractional precipitation followed by chromatography on DEAE 52‐cellulose column and then on Sephadex G‐75 column. The chitinase has a molecular weight of ca. 30 kDa as measured by sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS‐PAGE) electrophoresis. Its optimum pH is 4.5, and its optimum temperature is 50 °C with colloidal chitin as a substrate. The enzyme is stable below 45 °C and in pH ranges between 4.5 and 5.5. It is activated by glucosamine, glucose, N‐acetylglucosamine, and metal ions including Ca2+, Fe2+, Fe3+, and Ni2+. It is inhibited by SDS, H2O2, ascorbic acid, Cu2+, Mg2+, Ba2+, Sn2+, Cr3+, and K+. With colloidal chitin as substrate, the Km and the Vmax of the chitinase are 4.28 mg/mL and 14.29 μg/(Min·mL), respectively, whereas the end products of the enzymatic hydrolysis are 14.33% monomer and 85.67% dimer of N‐acetylglucosamine. The viscosity of carboxymethyl chitin decreased rapidly at the initial stages when subjected to chitinase hydrolysis, which indicates that the chitinase acts in an endosplitting pattern.