ABSTRACT
Chitosan-degrading activities induced by glucosamine (GlcN) orN-acetylglucosamine (GlcNAc) were found in a culture filtrate of Trichoderma reesei PC-3-7. One of the chitosan-degrading enzymes was purified to homogeneity by precipitation with ammonium sulfate followed by anion-exchange and hydrophobic-interaction chromatographies. The enzyme was monomeric, and its molecular mass was 93 kDa. The optimum pH and temperature of the enzyme were 4.0 and 50°C, respectively. The activity was stable in the pH range 6.0 to 9.0 and at a temperature below 50°C. Reaction product analysis from the viscosimetric assay and thin-layer chromatography and 1H nuclear magnetic resonance spectroscopy clearly indicated that the enzyme was an exo-type chitosanase, exo-β-d-glucosaminidase, that releases GlcN from the nonreducing end of the chitosan chain. 1H nuclear magnetic resonance spectroscopy also showed that the exo-β-d-glucosaminidase produced a β-form of GlcN, demonstrating that the enzyme is a retaining glycanase. Time-dependent liberation of the reducing sugar from partially acetylated chitosan with exo-β-d-glucosaminidase and the partially purified exo-β-d-N-acetylglucosaminidase from T. reesei PC-3-7 suggested that the exo-β-d-glucosaminidase cleaves the glycosidic link of either GlcN-β(1→4)-GlcN or GlcN-β(1→4)-GlcNAc.
Total and Class-Specific Analysis of Per- and Polyfluoroalkyl Substances in Environmental Samples Using Nuclear Magnetic Resonance Spectroscopy
