PURIFICATION AND SOME PROPERTIES OF THE EXTRACELLULAR CHITINASE PRODUCED BY TRICHODERMA-HARZIANUM

Chitinase (EC 3.2.1.14), from the culture filtrate of Trichoderma harzianum, was successively purified by precipitation with ammonium sulfate followed by ion-exchange chromatography on Q-Sepharose, gel filtration on Sephadex G-100, and hydrophobic interaction on Phenyl-Sepharose CL-4B. A typical procedure provided 76-fold purification with 10% yield. The molecular weight of the purified chitinase was found to be approximately 40,000 Da by SDS-PAGE and 36,000 Da by gel filtration. The pH optimum for the enzyme was pH 4.0-4.5, and maximum activity was obtained at 40-degrees-C. The chitinase rapidly hydrolysed swollen chitin and regenerated chitin, but glycol chitin and powdered chitin very slowly. N',N-Diacetylchitobiose was not affected by enzyme.