title: Crystal structure of grape dihydroflavonol 4-reductase, a key enzyme in flavonoid biosynthesis.
abstract: The nicotinamide adenine dinucleotide phosphate (NADPH)-dependent enzyme dihydroflavonol 4-reductase (DFR) catalyzes a late step in the biosynthesis of anthocyanins and condensed tannins, two flavonoid classes of importance to plant survival and human nutrition. This enzyme has been widely investigated in many plant species, but little is known about its structural and biochemical properties. To provide a basis for detailed structure-function studies, the crystal structure of Vitis vinifera DFR, heterologously expressed in Escherichia coli, has been determined at 1.8 A resolution. The 3D structure of the ternary complex obtained with the oxidized form of nicotinamide adenine dinucleotide phosphate and dihydroquercetin, one of the DFR substrates, presents common features with the short-chain dehydrogenase/reductase family, i.e., an N-terminal domain adopting a Rossmann fold and a variable C-terminal domain, which participates in substrate binding. The structure confirms the importance of the 131-156 region, which lines the substrate binding site and enlightens the role of a specific residue at position 133 (Asn or Asp), assumed to control substrate recognition. The activity of the wild-type enzyme and its variant N133D has been quantified in vitro, using dihydroquercetin or dihydrokaempferol. Our results demonstrate that position 133 cannot be solely responsible for the recognition of the B-ring hydroxylation pattern of dihydroflavonols.
authors: Chimie et Biologie des Membranes et des Nanoobjets, UMR CNRS 5248, Bâtiment B8, Avenue des Facultés, Université Bordeaux 1, 33405 Talence Cedex, France.
authors: Pierre
authors: P
authors: Petit
authors: Thierry
authors: T
authors: Granier
authors: Béatrice Langlois
authors: BL
authors: d'Estaintot
authors: Claude
authors: C
authors: Manigand
authors: Katell
authors: K
authors: Bathany
authors: Jean-Marie
authors: JM
authors: Schmitter
authors: Virginie
authors: V
authors: Lauvergeat
authors: Saïd
authors: S
authors: Hamdi
authors: Bernard
authors: B
authors: Gallois
mesh_terms: D000429:Alcohol Oxidoreductases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D005419:Flavonoids / Q000096:biosynthesis*; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D015394:Molecular Structure; D009249:NADP / Q000737:chemistry / Q000378:metabolism; D010084:Oxidation-Reduction; D010940:Plant Proteins / Q000737:chemistry* / Q000378:metabolism; D017434:Protein Structure, Tertiary; D027843:Vitis / Q000201:enzymology*
chemical_list: D005419:Flavonoids; D010940:Plant Proteins; D009249:NADP; D000429:Alcohol Oxidoreductases; C056570:dihydroflavanol 4-reductase