title: Biochemical characterization of the first fungal glycoside hydrolyase family 3 β-N-acetylglucosaminidase from Rhizomucor miehei.
abstract: A novel β-N-acetylglucosaminidase gene (RmNag) from Rhizomucor miehei was cloned and expressed in Escherichia coli. RmNag shares the highest identity of 37% with a putative β-N-acetylglucosaminidase from Aspergillus clavatus. The recombinant enzyme was purified to homogeneity. The optimal pH and temperature of RmNag were pH 6.5 and 50 °C, respectively. It was stable in the pH range 6.0-8.0 and at temperatures below 45 °C. RmNag exhibited strict substrate specificity for p-nitrophenyl β-N-acetylglucosaminide (pNP-GlcNAc) and N-acetyl chitooligosaccharides. The apparent Km of RmNag toward pNP-GlcNAc was 0.13 mM. The purified enzyme displayed an exo-type manner as it released the only end product of GlcNAc from all the tested N-acetyl chitooligosaccharides. Besides, RmNag exhibited relatively high N-acetyl-β-D-glucosaminide tolerance with an inhibition constant Ki value of 9.68 mM. The excellent properties may give the enzyme great potential in industries. This is the first report on a glycoside hydrolyase family 3 β-N-acetylglucosaminidase from a fungus.
authors.affiliation: Department of Biotechnology, College of Food Science and Nutritional Engineering, and ‡Bioresource Utilization Laboratory, College of Engineering, China Agricultural University , Beijing 100083, China.
authors.forename: Shaoqing
authors.initials: S
authors.lastname: Yang
authors.forename: Shuang
authors.initials: S
authors.lastname: Song
authors.forename: Qiaojuan
authors.initials: Q
authors.lastname: Yan
authors.forename: Xing
authors.initials: X
authors.lastname: Fu
authors.forename: Zhengqiang
authors.initials: Z
authors.lastname: Jiang
authors.forename: Xinbin
authors.initials: X
authors.lastname: Yang
mesh_terms: D000118:Acetylglucosaminidase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D004795:Enzyme Stability; D005656:Fungal Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006026:Glycoside Hydrolases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D007700:Kinetics; D008969:Molecular Sequence Data; D020103:Rhizomucor / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics; D016415:Sequence Alignment; D013379:Substrate Specificity
chemical_list: D005656:Fungal Proteins; D006026:Glycoside Hydrolases; D000118:Acetylglucosaminidase