title: Enzymatic synthesis of a neoglycoconjugate by transglycosylation with Arthrobacter endo-beta-N-acetylglucosaminidase: a substrate for colorimetric detection of endo-beta-N-acetylglucosaminidase activity.
abstract: The transglycosylation activity of endo-beta-N-acetylglucosaminidase from Arthrobacter protophormiae was used for the enzymatic synthesis of a novel oligosaccharide, Man6GlcNAc-p-nitrophenyl-alpha-D-glucose (Man6GlcNAc-Glc-pNP). The reaction was efficiently induced in aqueous solution containing dimethyl sulfoxide. In the medium containing 20% (v/v) dimethyl sulfoxide with 0.1 M Glc-pNP as an acceptor, the transglycosylation attained yields of 75% by high-performance anion-exchange chromatography. The structure of Man6GlcNAc-Glc-pNP was confirmed by ion mass spectrometry and 400 MHz 1H NMR spectrometry. Various endo-beta-N-acetylglucosaminidases hydrolyzed this oligosaccharide and Man6GlcNAc and Glc-pNP were released from the oligosaccharide by endo-beta-N-acetylglucosaminidase digestion. We have established a new procedure for the colorimetric detection of endo-beta-N-acetylglucosaminidase activity using Man6Glc-NAc-Glc-pNP, which is simple as that for other exoglycosidase assays with pNP-glycosides as substrates.
authors.affiliation: Department of Bioresource Science, Faculty of Agriculture, Kagawa University, Japan.
authors.forename: K
authors.initials: K
authors.lastname: Takegawa
authors.forename: K
authors.initials: K
authors.lastname: Fujita
authors.forename: J Q
authors.initials: JQ
authors.lastname: Fan
authors.forename: M
authors.initials: M
authors.lastname: Tabuchi
authors.forename: N
authors.initials: N
authors.lastname: Tanaka
authors.forename: A
authors.initials: A
authors.lastname: Kondo
authors.forename: H
authors.initials: H
authors.lastname: Iwamoto
authors.forename: I
authors.initials: I
authors.lastname: Kato
authors.forename: Y C
authors.initials: YC
authors.lastname: Lee
authors.forename: S
authors.initials: S
authors.lastname: Iwahara
mesh_terms: D000118:Acetylglucosaminidase / Q000737:chemistry / Q000378:metabolism*; D001173:Arthrobacter / Q000201:enzymology*; D002240:Carbohydrate Sequence; D003124:Colorimetry; D004121:Dimethyl Sulfoxide; D005417:Flavobacterium / Q000201:enzymology; D006031:Glycosylation; D006868:Hydrolysis; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000138:chemical synthesis / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013302:Streptomyces / Q000201:enzymology; D015003:Yeasts / Q000201:enzymology
chemical_list: D009844:Oligosaccharides; D011994:Recombinant Proteins; C112131:mannosyl(6)-N-acetylglucosamine-p-nitrophenyl-alpha-D-glucose; D000118:Acetylglucosaminidase; D004121:Dimethyl Sulfoxide