Chitooligosaccharides (COS) generated from either chitin (chitin oligosaccharides) or chitosan (chitosan oligosaccharides) have a wide range of applications in agriculture, medicine, and other fields. Here, we report the characterization of a chitosanase from Bacillus amyloliquefaciens (BamCsn) and the importance of a tryptophan (Trp), W204, for BamCsn activity. BamCsn hydrolyzed the chitosan polymer by an endo mode. It also hydrolyzed chitin oligosaccharides and interestingly exhibited transglycosylation activity on chitotetraose and chitopentaose. Mutation of W204, a nonconserved amino acid in chitosanases, to W204A abolished the hydrolytic activity of BamCsn, with a change in the structure that resulted in a decreased affinity for the substrate and impaired the catalytic ability. Phylogenetic analysis revealed that BamCsn could belong to a new class of chitosanases that showed unique properties like transglycosylation, cleavage of chitin oligosaccharides, and the presence of W204 residues, which is important for activity. Chitosanases belonging to the BamCsn class showed a high potential to generate COS from chitinous substrates.
Keywords: BamCsn; chitooligosaccharides; differential scanning calorimetry; isothermal titration calorimetry; transglycosylation.