title: Structural and biochemical characterization of a trapped coenzyme A adduct of Caenorhabditis elegans glucosamine-6-phosphate N-acetyltransferase 1.
abstract: Glucosamine-6-phosphate N-acetyltransferase 1 (GNA1) produces GlcNAc-6-phosphate from GlcN-6-phosphate and acetyl coenzyme A. Early mercury-labelling experiments implicated a conserved cysteine in the reaction mechanism, whereas recent structural data appear to support a mechanism in which this cysteine plays no role. Here, two crystal structures of Caenorhabditis elegans GNA1 are reported, revealing an unusual covalent complex between this cysteine and the coenzyme A product. Mass-spectrometric and reduction studies showed that this inactive covalent complex can be reactivated through reduction, yet mutagenesis of the cysteine supports a previously reported bi-bi mechanism. The data unify the apparently contradictory earlier reports on the role of a cysteine in the GNA1 active site.
authors.affiliation: Division of Molecular Microbiology, College of Life Sciences, University of Dundee, Dundee DD1 5EH, Scotland.
authors.forename: Helge C
authors.initials: HC
authors.lastname: Dorfmueller
authors.forename: Wenxia
authors.initials: W
authors.lastname: Fang
authors.forename: Francesco V
authors.initials: FV
authors.lastname: Rao
authors.forename: David E
authors.initials: DE
authors.lastname: Blair
authors.forename: Helen
authors.initials: H
authors.lastname: Attrill
authors.forename: Daan M F
authors.initials: DM
authors.lastname: van Aalten
mesh_terms: D000818:Animals; D017173:Caenorhabditis elegans / Q000201:enzymology*; D020134:Catalytic Domain; D003001:Cloning, Molecular; D003065:Coenzyme A / Q000737:chemistry*; D017124:Conserved Sequence; D018360:Crystallography, X-Ray / Q000379:methods; D003545:Cysteine / Q000737:chemistry; D051081:Glucosamine 6-Phosphate N-Acetyltransferase / Q000737:chemistry*; D007700:Kinetics; D013058:Mass Spectrometry / Q000379:methods; D008958:Models, Molecular; D008968:Molecular Conformation; D009154:Mutation; D010100:Oxygen / Q000737:chemistry; D011485:Protein Binding
chemical_list: D051081:Glucosamine 6-Phosphate N-Acetyltransferase; D003545:Cysteine; D010100:Oxygen; D003065:Coenzyme A