title: The N-acetyl-binding pocket of N-acetylglucosaminyltransferases also accommodates a sugar analog with a chemical handle at C2.
abstract: In recent years, sugars with a unique chemical handle have been used to detect and elucidate the function of glycoconjugates. Such chemical handles have generally been part of an N-acetyl moiety of a sugar. We have previously developed several applications using the single mutant Y289L-β1,4-galactosyltransferase I (Y289L-β4Gal-T1) and the wild-type polypeptide-α-GalNAc-T enzymes with UDP-C2-keto-Gal. Here, we describe for the first time that the GlcNAc-transferring enzymes-R228K-Y289L-β4Gal-T1 mutant enzyme, the wild-type human β1,3-N-acetylglucosaminyltransferase-2 and human Maniac Fringe-can also transfer the GlcNAc analog C2-keto-Glc molecule from UDP-C2-keto-Glc to their respective acceptor substrates. Although the R228K-Y289L-β4Gal-T1 mutant enzyme transfers the donor sugar substrate GlcNAc or its analog C2-keto-Glc only to its natural acceptor substrate, GlcNAc, it does not transfer to its analog C2-keto-Glc. Thus, these observations suggest that the GlcNAc-transferring glycosyltransferases can generally accommodate a chemical handle in the N-acetyl-binding cavity of the donor sugar substrate, but not in the N-acetyl-binding cavity of the acceptor sugar.
authors.affiliation: Structural Glycobiology Section, Center for Cancer Research Nanobiology Program, Frederick, MD 21702, USA.
authors.forename: Marta
authors.initials: M
authors.lastname: Pasek
authors.forename: Boopathy
authors.initials: B
authors.lastname: Ramakrishnan
authors.forename: Elizabeth
authors.initials: E
authors.lastname: Boeggeman
authors.forename: Natalia
authors.initials: N
authors.lastname: Mercer
authors.forename: Andres E
authors.initials: AE
authors.lastname: Dulcey
authors.forename: Gary L
authors.initials: GL
authors.lastname: Griffiths
authors.forename: Pradman K
authors.initials: PK
authors.lastname: Qasba
mesh_terms: D000117:Acetylglucosamine / Q000737:chemistry; D019943:Amino Acid Substitution; D001665:Binding Sites; D002236:Carbohydrate Conformation; D002240:Carbohydrate Sequence; D002846:Chromatography, Affinity; D003001:Cloning, Molecular; D004926:Escherichia coli; D005167:Factor VII / Q000737:chemistry; D005690:Galactose / Q000031:analogs & derivatives* / Q000737:chemistry*; D005964:Glucosyltransferases; D006031:Glycosylation; D006602:Hexosyltransferases / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D006801:Humans; D006860:Hydrogen Bonding; D057927:Hydrophobic and Hydrophilic Interactions; D047908:Intracellular Signaling Peptides and Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D008565:Membrane Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D008958:Models, Molecular; D008969:Molecular Sequence Data; D050505:Mutant Proteins / Q000737:chemistry; D017351:N-Acetylglucosaminyltransferases / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D009844:Oligosaccharides / Q000737:chemistry; D011485:Protein Binding; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D013499:Surface Properties
chemical_list: D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D050505:Mutant Proteins; D009844:Oligosaccharides; D011993:Recombinant Fusion Proteins; D005167:Factor VII; D005964:Glucosyltransferases; D006602:Hexosyltransferases; D017351:N-Acetylglucosaminyltransferases; C106516:MFNG protein, human; D000117:Acetylglucosamine; D005690:Galactose