title: The active site of spinach glycolate oxidase.
abstract: The amino acid sequence of glycolate oxidase from spinach has been fitted to an electron density map at 2.2 A resolution. From a refined model we give a detailed description of the flavin mononucleotide-binding site and the residues which might be involved in the catalytic action of the enzyme. The cofactor is bound to the enzyme at the carboxy end of the beta strands in the alpha/beta barrel domain and forms a number of hydrogen bonds both to residues at the end of the beta strands and in the loop regions after the strands. In particular Lys-230 interacts with atoms N1 and O2 of the isoalloxazine ring, which produces an inductive effect that could enhance the nucleophilicity of the electron acceptor N5 of the flavin ring. Almost the entire coenzyme is buried in the interior of the enzyme. The exceptions are one phosphate oxygen atom and a region around the N5 position on the si-face of the isoalloxazine ring, which are accessible to solution. Based on a model of bound glycolate to our structure we propose that the following residues are important in the catalytic reaction: Arg-257, Tyr-24, and Tyr-129 for binding the substrate, and His-254 for abstracting a proton from the C2 atom of the substrate.
authors.affiliation: Department of Molecular Biology, Swedish University of Agricultural Sciences, Uppsala Biomedical Center.
authors.forename: Y
authors.initials: Y
authors.lastname: Lindqvist
authors.forename: C I
authors.initials: CI
authors.lastname: Brändén
mesh_terms: D000429:Alcohol Oxidoreductases / Q000378:metabolism; D000595:Amino Acid Sequence; D001665:Binding Sites; D055598:Chemical Phenomena; D002621:Chemistry; D005486:Flavin Mononucleotide / Q000378:metabolism; D006860:Hydrogen Bonding; D046911:Macromolecular Substances; D008958:Models, Molecular; D015394:Molecular Structure; D009584:Nitrogen; D010100:Oxygen; D010944:Plants / Q000201:enzymology*; D011487:Protein Conformation; D014675:Vegetables
chemical_list: D046911:Macromolecular Substances; D005486:Flavin Mononucleotide; D000429:Alcohol Oxidoreductases; C020821:glycollate oxidase; D009584:Nitrogen; D010100:Oxygen