title: Serratia marcescens chitobiase is a retaining glycosidase utilizing substrate acetamido group participation.
abstract: The stereochemistry of the reaction catalysed by Serratia marcescens chitobiase was determined by HPLC separation of the anomers of N-acetylglucosamine produced during the hydrolysis of p-nitrophenyl N-acetyl-beta-d-glucosaminide (PNP-GlcNAc). In the early stages of the reaction, the beta-anomer was found to prevail, whereas the alpha-anomer dominated at mutarotation equilibrium. This established that chitobiase hydrolyses glycosidic bonds with overall retention of the anomeric configuration. Chitobiase-catalysed hydrolysis of PNP-GlcNAc was competitively inhibited by a series of chito-oligosaccharides (degree of polymerization 2-5) that were selectively de-N-acetylated at their non-reducing end. The results are in accord with the participation of the acetamido group at C-2 of the substrate in the catalytic mechanism of chitobiase and related enzymes.
authors.affiliation: Centre de Recherches sur les Macromolécules Végétales, C.N.R.S., BP 53, F-38041 Grenoble cedex 9, France.
authors.forename: S
authors.initials: S
authors.lastname: Drouillard
authors.forename: S
authors.initials: S
authors.lastname: Armand
authors.forename: G J
authors.initials: GJ
authors.lastname: Davies
authors.forename: C E
authors.initials: CE
authors.lastname: Vorgias
authors.forename: B
authors.initials: B
authors.lastname: Henrissat
mesh_terms: D000107:Acetylation; D000117:Acetylglucosamine / Q000031:analogs & derivatives / Q000737:chemistry / Q000378:metabolism; D000118:Acetylglucosaminidase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D002240:Carbohydrate Sequence; D002851:Chromatography, High Pressure Liquid; D004791:Enzyme Inhibitors; D004926:Escherichia coli / Q000235:genetics; D006026:Glycoside Hydrolases / Q000378:metabolism; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000737:chemistry / Q000494:pharmacology; D011994:Recombinant Proteins / Q000378:metabolism; D012706:Serratia marcescens / Q000201:enzymology*; D013237:Stereoisomerism
chemical_list: D004791:Enzyme Inhibitors; D009844:Oligosaccharides; D011994:Recombinant Proteins; C014124:4-nitrophenyl-N-acetyl-2-deoxyglucopyranoside; D006026:Glycoside Hydrolases; D000118:Acetylglucosaminidase; D000117:Acetylglucosamine