Chitinases of Streptomyces kurssanovii: purification and some properties

Four major chitinases, with molecular weights of 42, 40, 26 and 20 kDa, were detected in the culture supernatant of Streptomyces kurssanovii. They were isolated by means of metal-ion affinity chromatography and hydrophobic-interaction chromatography on butyl-toyopearl 650 M and octyl-sepharose CL-4B with a recovery of 57% of total initial activity. The purified chitinases were homogeneous according to SDS-PAGE. For two enzymes, Chi-42 and Chi-26, molecular weights, pI values, amino acid composition, as well as the influence of pH, temperature and some metal ions on activities and stabilities were determined. The chitinases Chi-42 and Chi-26 hydrolysed chitin, chitosan (85% deacetylated) and CM-chitin (69% substituted) in an endo-splitting manner. The apparent Km values for Chi-42 calculated from Lineweaver-Burk plots were 0·16 × 10−4 M (chitosan) and 0·18 × 10−4 M (CM-chitin) at pH = 4·0 and t = 37°C. Both chitinases hydrolysed tetramers and pentamers of N-acetylglucosamine, but less readily hydrolysed its trimer. The HPLC analysis of the reaction products indicated that the chitinase Chi-26 also catalysed a transglycosylation reaction, since oligosaccharides with a higher degree of polymerization than the initial substrates were produced.