Structure and epimerase activity of anthocyanidin reductase from Vitis vinifera.
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 A and explains the inactivity of the enzyme in the presence of high salt concentrations.
Gargouri
Mahmoud
M
Chimie et Biologie des Macromolécules et des Nanoobjets UMR CNRS 5248, Bâtiment B8, Avenue des Facultés, Université Bordeaux 1, Talence CEDEX, France.
Manigand
Claude
C
Maugé
Chloé
C
Granier
Thierry
T
Langlois d'Estaintot
Béatrice
B
Cala
Olivier
O
Pianet
Isabelle
I
Bathany
Katell
K
Chaudière
Jean
J
Gallois
Bernard
B
D000494:Allosteric Regulation; D000872:Anthocyanins / Q000378:metabolism*; D003460:Crystallization; D018360:Crystallography, X-Ray; D004926:Escherichia coli / Q000235:genetics; D007536:Isomerism; D009247:NADH, NADPH Oxidoreductases / Q000737:chemistry* / Q000235:genetics; D010084:Oxidation-Reduction; D011487:Protein Conformation; D019998:Racemases and Epimerases / Q000737:chemistry* / Q000235:genetics; D013329:Structure-Activity Relationship; D019076:Transgenes / Q000235:genetics; D027843:Vitis / Q000201:enzymology
D000872:Anthocyanins; D009247:NADH, NADPH Oxidoreductases; D019998:Racemases and Epimerases