Enrichment of chitinolytic microorganisms:: isolation and characterization of a chitinase exhibiting antifungal activity against phytopathogenic fungi from a novel Streptomyces strain

Thirteen different chitin-degrading bacteria were isolated from soil and sediment samples. Five of these strains (SGE2, SGE4, SSL3, MG1, and MG3) exhibited antifungal activity against phytopathogenic fungi. Analyses of the 16S rRNA genes and the substrate spectra revealed that the isolates belong to the genera Bacillus or Streptomyces. The closest relatives were Bacillus chitinolyticus ( SGE2, SGE4, and SSL3), B. ehimensis ( MG1), and Streptomyces griseus ( MG3). The chitinases present in the culture supernatants of the five isolates revealed optimal activity between 45 degreesC and 50 degreesC and at pH values of 4 ( SSL3), 5 ( SGE2 and MG1), 6 ( SGE4), and 5 - 7 ( MG3). The crude chitinase preparations of all five strains possessed antifungal activity. The chitinase of MG3 (ChiIS) was studied further, since the crude enzyme conferred strong growth suppression of all fungi tested and was very active over the entire pH range tested. The chiIS gene was cloned and the gene product was purified. The deduced protein consisted of 303 amino acids with a predicted molecular mass of 31,836 Da. Sequence analysis revealed that ChiIS of MG3 is similar to chitinases of Streptomyces species, which belong to family 19 of glycosyl hydrolases. Purified ChiIS showed remarkable antifungal activity and stability.