title: Purification and properties of extracellular chitinases from the parasitic fungus Isaria japonica.
abstract: Two chitinases (P-1 and P-2) induced with colloidal chitin were purified from the culture supernatant of Isaria japonica by chromatography on DEAE Bio-Gel, chromatofocusing and gel filtration with Superdex 75 pg. The enzymes were electrophoretically homogeneous and estimated to have a molecular mass of 43,273 (+/-5) for P-1 and 31,134 (+/-6) for P-2 by MALDI-MS. The optimum pH and temperature was 3.5-4.0 and 50 degrees C for P-1 and 4.0-4.5 and 40 degrees C for P-2. P-1 acted against chitosan 7B (degree of deacetylation, 65-74%) = glycol chitin > colloidal chitin = chitosan 10B (degree of deacetylation, above 99%) and P-2 against chitosan 7B > glycol chitin = chitosan 10B > colloidal chitin in order of activity. The products of hydrolysis of chitin and chitosan hexamer were analyzed by MALDI-MS. The products from the chitin hexamer obtained with P-1 were almost all dimers with only a small amount of trimer whereas those obtained with P-2 were mainly trimers with some dimer and tetramer. No hydrolysis of chitosan hexamer was observed. High homology in the amino-terminal sequence for chitinase P-1 was exhibited by chitinases from Trichoderma harzianum, Candida albicans and Saccharomyces cerevisiae in the range of 48-39%. The highest homology for Chitinase P-2 was shown by an endochitinase from Metarhizium anisopliae of 66%, while 44% homology was exhibited by chitinases of Leguminosae plants.
authors.affiliation: Faculty of Agriculture, Laboratory of Biological Chemistry, Meijo University, 1-501 Shiogamaguchi, Tempaku-ku, Nagoya 468-8502, Japan.
authors.forename: I
authors.initials: I
authors.lastname: Kawachi
authors.forename: T
authors.initials: T
authors.lastname: Fujieda
authors.forename: M
authors.initials: M
authors.lastname: Ujita
authors.forename: Y
authors.initials: Y
authors.lastname: Ishii
authors.forename: K
authors.initials: K
authors.lastname: Yamagishi
authors.forename: H
authors.initials: H
authors.lastname: Sato
authors.forename: T
authors.initials: T
authors.lastname: Funaguma
authors.forename: A
authors.initials: A
authors.lastname: Hara