Isolation and functional characterization of a cDNA coding a hydroxycinnamoyltransferase involved in phenylpropanoid biosynthesis in Cynara cardunculus L.
Isolation and functional characterization of a cDNA coding a hydroxycinnamoyltransferase involved in phenylpropanoid biosynthesis in Cynara cardunculus L.
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DOI：
10.1186/1471-2229-7-14
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发表时间：
2007-03-20
影响因子：
5.3
通讯作者：
Bourgaud F
中科院分区：
文献类型：
--
作者：
Comino C;Lanteri S;Portis E;Acquadro A;Romani A;Hehn A;Larbat R;Bourgaud F
Cynara cardunculus L.是一种具有药用价值的食用植物，特别是在其叶子的多酚含量方面。它包括三个分类群：球蓟、栽培豆角和野生豆角。主要的酚类化合物是二咖啡酰奎宁酸(如野百合)，它们主要局限于Cynara物种，以及它们的前体绿原酸(CGA)。这项研究的范围是为了更好地了解这种植物中CGA的合成。鉴定了一个编码与CGA合成有关的羟基肉桂酰基转移酶(HCT)的基因序列。基因序列的分离是通过使用针对同源hct序列保守区的简并引物的聚合酶链式反应策略实现的。我们克隆了一个717个碱基的cDNA，其氨基酸同源性为84%，与一个由对香豆酰辅酶A和奎尼酸生物合成CGA的烟草基因有92%的相似性。在计算机研究中发现，朝鲜蓟Hct序列与一个主要的酰基转移酶(即邻氨基苯甲酸N-羟基肉桂酰基/苯甲酰基转移酶)聚在一起。在大肠杆菌中异源表达了全长的HCT(GenBank AccessionDQ104740)，证明该重组酶能有效地从奎尼酸和咖啡酰辅酶A或对香豆酰辅酶A合成绿原酸和对香豆酰奎宁酸酯，证实其为羟基肉桂酰辅酶A：奎宁酸酯。野生和栽培的Cardunculus亚种之间表现出不同水平的Hct表达。表达水平与CGA含量呈正相关。这些数据支持Cynara cardunculus Hct在羟基肉桂酸酯羟基化步骤之前和/或之后参与CGA生物合成的预测。
Cynara cardunculus L. is an edible plant of pharmaceutical interest, in particular with respect to the polyphenolic content of its leaves. It includes three taxa: globe artichoke, cultivated cardoon, and wild cardoon. The dominating phenolics are the di-caffeoylquinic acids (such as cynarin), which are largely restricted to Cynara species, along with their precursor, chlorogenic acid (CGA). The scope of this study is to better understand CGA synthesis in this plant. A gene sequence encoding a hydroxycinnamoyltransferase (HCT) involved in the synthesis of CGA, was identified. Isolation of the gene sequence was achieved by using a PCR strategy with degenerated primers targeted to conserved regions of orthologous HCT sequences available. We have isolated a 717 bp cDNA which shares 84% aminoacid identity and 92% similarity with a tobacco gene responsible for the biosynthesis of CGA from p-coumaroyl-CoA and quinic acid. In silico studies revealed the globe artichoke HCT sequence clustering with one of the main acyltransferase groups (i.e. anthranilate N-hydroxycinnamoyl/benzoyltransferase). Heterologous expression of the full length HCT (GenBank accession DQ104740) cDNA in E. coli demonstrated that the recombinant enzyme efficiently synthesizes both chlorogenic acid and p-coumaroyl quinate from quinic acid and caffeoyl-CoA or p-coumaroyl-CoA, respectively, confirming its identity as a hydroxycinnamoyl-CoA: quinate HCT. Variable levels of HCT expression were shown among wild and cultivated forms of C. cardunculus subspecies. The level of expression was correlated with CGA content. The data support the predicted involvement of the Cynara cardunculus HCT in the biosynthesis of CGA before and/or after the hydroxylation step of hydroxycinnamoyl esters.
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影响因子：
46.9
作者：
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通讯作者：
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