title: Catalytic function of a newly purified exo-β-D-glucosaminidase from the entomopathogenic fungus Paecilomyces lilacinus.
abstract: An entomopathogenic fungus, Paecilomyces lilacinus, was found to grow on chitosanase-detecting plates. Besides an endo-chitosanase, an exo-β-D-glucosaminidase was purified by cation-exchange chromatography from this microorganism cultivated in M9 minimal media containing 0.5% chitosan as the sole carbon source. The molecular weight of the enzyme is 95kDa; the optimum pH and temperature for activity are 6.0 and 45°C, respectively. The purified exo-β-D-GlcNase promotes the hydrolysis of 95% deacetylated chitosan from its non-reducing end and liberates 2-amino-2-deoxy-D-glucopyranose (GlcN) as the sole product; however, 2-acetamido-2-deoxy-D-glucopyranose (GlcNAc) was not detected when chitin was used as the substrate. The cleavage pattern confirmed using real-time mass spectrometry shows that exo-β-D-glucosaminidase cleaves the glycosidic bonds between GlcN-GlcN and GlcN-GlcNAc but not between GlcNAc-GlcN or GlcNAc-GlcNAc. In the presence of a 10% solution of various alcohols, many alkyl-β-D-glucosaminides were obtained, indicating that exo-β-D-glucosaminidase is a retaining enzyme.
authors.affiliation: Department of Applied Chemistry, National Chiao Tung University, 1001 Ta-Hseh Road, HsinChu, Taiwan.
authors.forename: Cheng-Fu
authors.initials: CF
authors.lastname: Chao
authors.forename: Yi-Yun
authors.initials: YY
authors.lastname: Chen
authors.forename: Chih-Yu
authors.initials: CY
authors.lastname: Cheng
authors.forename: Yaw-Kuen
authors.initials: YK
authors.lastname: Li
mesh_terms: D000107:Acetylation; D002686:Chitin / Q000378:metabolism; D048271:Chitosan / Q000378:metabolism; D003470:Culture Media / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation; D004790:Enzyme Induction; D004795:Enzyme Stability; D005656:Fungal Proteins / Q000096:biosynthesis / Q000302:isolation & purification*; D006031:Glycosylation; D006596:Hexosaminidases / Q000096:biosynthesis / Q000302:isolation & purification*; D006868:Hydrolysis; D025282:Mycelium / Q000201:enzymology; D010143:Paecilomyces / Q000201:enzymology*; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods; D013237:Stereoisomerism; D013379:Substrate Specificity
chemical_list: D003470:Culture Media; D005656:Fungal Proteins; D002686:Chitin; D048271:Chitosan; D006596:Hexosaminidases; C064499:exo-beta-D-glucosaminidase