Abstract
Catechins (flavan-3-ols), the most important secondary metabolites in the tea plant, have positive effects on human health and are crucial in defense against pathogens of the tea plant. The aim of this study was to elucidate the biosynthetic pathway of galloylated catechins in the tea plant. The results suggested that galloylated catechins were biosynthesized via 1-O-glucose ester-dependent two-step reactions by acyltransferases, which involved two enzymes, UDP-glucose:galloyl-1-O-β-d-glucosyltransferase (UGGT) and a newly discovered enzyme, epicatechin:1-O-galloyl-β-d-glucose O-galloyltransferase (ECGT). In the first reaction, the galloylated acyl donor β-glucogallin was biosynthesized by UGGT from gallic acid and uridine diphosphate glucose. In the second reaction, galloylated catechins were produced by ECGT catalysis from β-glucogallin and 2,3-cis-flavan-3-ol. 2,3-cis-Flavan-3-ol and 1-O-galloyl-β-d-glucose were appropriate substrates of ECGT rather than 2,3-trans-flavan-3-ol and 1,2,3,4,6-pentagalloylglucose. Purification by more than 1641-fold to apparent homogeneity yielded ECGT with an estimated molecular mass of 241 to 121 kDa by gel filtration. Enzyme activity and SDS-PAGE analysis indicated that the native ECGT might be a dimer, trimer, or tetramer of 60- and/or 58-kDa monomers, and these monomers represent a heterodimer consisting of pairs of 36- or 34- of and 28-kDa subunits. MALDI-TOF-TOF MS showed that the protein SCPL1199 was identified. Epigallocatechin and epicatechin exhibited higher substrate affinities than β-glucogallin. ECGT had an optimum temperature of 30 °C and maximal reaction rates between pH 4.0 and 6.0. The enzyme reaction was inhibited dramatically by phenylmethylsulfonyl fluoride, HgCl2, and sodium deoxycholate.
Introduction
Flavonoids, a major class of secondary metabolites in plants, have a number of important physiological roles as endogenous auxin transport regulators, root development , seed germination, allelopathy , plant-bacterium interaction , UV-B protection , and plant defense against pathogens and environmental stress.
Flavonoids can be grouped into several subgroups including chalcone, flavone, flavonol, flavandiol, anthocyanin, proanthocyanidin (oligomer or polymer of flavan-3-ols and flavan-3,4-diol units) and other specialized forms. Flavan-3-ols (catechins), which comprise ∼70–80% of tea polyphenols, are rich in young leaves and shoots of the tea plant (Camellia sinensis (L.) O. Kuntze). Catechins, with a basic 2-phenylchromone structure, are characterized by the di- or tri-hydroxyl group substitution of the B ring, the 2,3-position isomer of the C ring, and presence of a galloyl group at the 3-postion of the C ring . On the basis of the classical definition proposed of galloyl group structural features, catechins are divided into galloylated and nongalloylated compounds. Galloylated catechins, including (−)-epigallocatechin gallate (EGCG)3 and (−)-epicatechin gallate (ECG), esterified often with gallic acid (GA) in the 3-hydroxyl group of the flavan-3-ol units are major catechin compounds that account for up to 76% of catechins in the tea plant .
Resource: http://www.ncbi.nlm.nih.gov
Read More: http://www.nutritionforest.com/
Catechins (flavan-3-ols), the most important secondary metabolites in the tea plant, have positive effects on human health and are crucial in defense against pathogens of the tea plant. The aim of this study was to elucidate the biosynthetic pathway of galloylated catechins in the tea plant. The results suggested that galloylated catechins were biosynthesized via 1-O-glucose ester-dependent two-step reactions by acyltransferases, which involved two enzymes, UDP-glucose:galloyl-1-O-β-d-glucosyltransferase (UGGT) and a newly discovered enzyme, epicatechin:1-O-galloyl-β-d-glucose O-galloyltransferase (ECGT). In the first reaction, the galloylated acyl donor β-glucogallin was biosynthesized by UGGT from gallic acid and uridine diphosphate glucose. In the second reaction, galloylated catechins were produced by ECGT catalysis from β-glucogallin and 2,3-cis-flavan-3-ol. 2,3-cis-Flavan-3-ol and 1-O-galloyl-β-d-glucose were appropriate substrates of ECGT rather than 2,3-trans-flavan-3-ol and 1,2,3,4,6-pentagalloylglucose. Purification by more than 1641-fold to apparent homogeneity yielded ECGT with an estimated molecular mass of 241 to 121 kDa by gel filtration. Enzyme activity and SDS-PAGE analysis indicated that the native ECGT might be a dimer, trimer, or tetramer of 60- and/or 58-kDa monomers, and these monomers represent a heterodimer consisting of pairs of 36- or 34- of and 28-kDa subunits. MALDI-TOF-TOF MS showed that the protein SCPL1199 was identified. Epigallocatechin and epicatechin exhibited higher substrate affinities than β-glucogallin. ECGT had an optimum temperature of 30 °C and maximal reaction rates between pH 4.0 and 6.0. The enzyme reaction was inhibited dramatically by phenylmethylsulfonyl fluoride, HgCl2, and sodium deoxycholate.
Introduction
Flavonoids, a major class of secondary metabolites in plants, have a number of important physiological roles as endogenous auxin transport regulators, root development , seed germination, allelopathy , plant-bacterium interaction , UV-B protection , and plant defense against pathogens and environmental stress.
Flavonoids can be grouped into several subgroups including chalcone, flavone, flavonol, flavandiol, anthocyanin, proanthocyanidin (oligomer or polymer of flavan-3-ols and flavan-3,4-diol units) and other specialized forms. Flavan-3-ols (catechins), which comprise ∼70–80% of tea polyphenols, are rich in young leaves and shoots of the tea plant (Camellia sinensis (L.) O. Kuntze). Catechins, with a basic 2-phenylchromone structure, are characterized by the di- or tri-hydroxyl group substitution of the B ring, the 2,3-position isomer of the C ring, and presence of a galloyl group at the 3-postion of the C ring . On the basis of the classical definition proposed of galloyl group structural features, catechins are divided into galloylated and nongalloylated compounds. Galloylated catechins, including (−)-epigallocatechin gallate (EGCG)3 and (−)-epicatechin gallate (ECG), esterified often with gallic acid (GA) in the 3-hydroxyl group of the flavan-3-ol units are major catechin compounds that account for up to 76% of catechins in the tea plant .
Resource: http://www.ncbi.nlm.nih.gov
Read More: http://www.nutritionforest.com/
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