Structure and epimerase activity of anthocyanidin reductase from Vitis vinifera

• Published in: Acta crystallographica. Section D, Biological crystallography. Biological crystallography., 2009-09, Vol.65 (9), p.989-1000
• Main Authors: Gargouri, Mahmoud, Manigand, Claude, Maugé, Chloé, Granier, Thierry, Langlois d'Estaintot, Béatrice, Cala, Olivier, Pianet, Isabelle, Bathany, Katell, Chaudière, Jean, Gallois, Bernard
• Format: Article
• Language: English
• Subjects: Allosteric Regulation; anthocyanidin reductases; Anthocyanins - metabolism; Crystallization; Crystallography, X-Ray; Escherichia coli; Escherichia coli - genetics; flavan-3-ols; flavonoid biosynthesis; Isomerism; NADH, NADPH Oxidoreductases - chemistry; NADH, NADPH Oxidoreductases - genetics; Oxidation-Reduction; Protein Conformation; Racemases and Epimerases - chemistry; Racemases and Epimerases - genetics; short-chain dehydrogenase reductases; Structure-Activity Relationship; Transgenes - genetics; Vitaceae; Vitis - enzymology; Vitis vinifera
• ISSN: 1399-0047; 0907-4449; 1399-0047
• DOI: 10.1107/S0907444909025013

Together with leucoanthocyanidin reductase, anthocyanidin reductase (ANR) is one of the two enzymes of the flavonoid‐biosynthesis pathway that produces the flavan‐3‐ol monomers required for the formation of proanthocyanidins or condensed tannins. It has been shown to catalyse the double reduction of anthocyanidins to form 2R,3R‐flavan‐3‐ols, which can be further transformed to the 2S,3R isomers by non‐enzymatic epimerization. ANR from grape (Vitis vinifera) was expressed in Escherichia coli and purified. Unexpectedly, RP‐HPLC, LC‐MS and NMR experiments clearly established that the enzyme produces a 50:50 mixture of 2,3‐cis and 2,3‐trans flavan‐3‐ols which have been identified by chiral chromatography to be 2S,3S‐ and 2S,3R‐flavan‐3‐ols, i.e. the naturally rare (+)‐epicatechin and (−)‐catechin, when cyanidin is used as the substrate of the reaction. The first three‐dimensional structure of ANR is described at a resolution of 2.2 Å and explains the inactivity of the enzyme in the presence of high salt concentrations.