Characterization and functional identification of a novel plant 4,5-extradiol dioxygenase involved in betalain pigment biosynthesis in Portulaca grandiflora
Characterization and functional identification of a novel plant 4,5-extradiol dioxygenase involved in betalain pigment biosynthesis in Portulaca grandiflora
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DOI：
10.1104/pp.103.031914
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发表时间：
2004-01-01
期刊：
影响因子：
7.4
通讯作者：
Zryd, JP
中科院分区：
文献类型：
--
作者：
Christinet, L;Burdet, FRX;Zryd, JP
甜菜素是一种色素，在石竹目植物的大多数家庭中取代花青素。甜菜碱酸是甜菜碱最常见的生色团。甜菜碱生物合成途径的关键酶是外切的双加氧酶，它打开碳4和碳5之间的二羟基苯丙氨酸(DOPA)的环，从而产生不稳定的Seco-DOPA，非酶重排为倍他氨酸。一个4，5-DOPA-双加氧酶的基因已经从真菌鹅膏中分离出来，但在植物中还没有发现同源基因。为了鉴定植物基因，我们构建了不同颜色表现型的马齿苋等基因系之间以及不同花芽形成阶段之间的消减文库。在差异表达的cDNA的电子分析中，我们发现了一个在翻译蛋白水平上与几种细菌外醇4，5-双加氧酶中存在的LigB结构域具有很强同源性的候选基因。该基因只在彩色花瓣中表达。该基因在甜菜碱生物合成途径中的功能被通过在缺乏颜色形成基因的大花杨的白色花瓣中进行生物遗传互补来证实。这个名为DodA的基因是一个新的植物双加氧酶家族的第一个特征成员，该家族在系统发育上与鹅膏菌不同。多巴双加氧酶。DODA的同源物不仅存在于甜菜碱产植物中，而且还存在于其他被子植物和苔藓植物中，尽管在催化部位附近有一些变化。这些同源基因是一个新的保守的植物基因家族的一部分，可能参与芳香化合物的代谢。
Betalains are pigments that replace anthocyanins in the majority of families of the plant order Caryophyllales. Betalamic acid is the common chromophore of betalains. The key enzyme of the betalain biosynthetic pathway is an extradiol dioxygenase that opens the cyclic ring of dihydroxy-phenylalanine (DOPA) between carbons 4 and 5, thus producing an unstable seco-DOPA that rearranges nonenzymatically to betalamic acid. A gene for a 4,5-DOPA-dioxygenase has already been isolated from the fungus Amanita muscaria, but no homolog was ever found in plants. To identify the plant gene, we constructed subtractive libraries between different colored phenotypes of isogenic lines of Portulaca grandiflora (Portulacaceae) and between different stages of flower bud formation. Using in silico analysis of differentially expressed cDNAs, we identified a candidate showing strong homology at the level of translated protein with the LigB domain present in several bacterial extradiol 4,5-dioxygenases. The gene was expressed only in colored flower petals. The function of this gene in the betalain biosynthetic pathway was confirmed by biolistic genetic complementation in white petals of P. grandiflora genotypes lacking the gene for color formation. This gene named DODA is the first characterized member of a novel family of plant dioxygenases phylogenetically distinct from Amanita sp. DOPA-dioxygenase. Homologs of DODA are present not only in betalain-producing plants but also, albeit with some changes near the catalytic site, in other angiosperms and in the bryophyte Physcomitrella patens. These homologs are part of a novel conserved plant gene family probably involved in aromatic compound metabolism.