Citrus plants have diverse methoxyflavonoids including, chrysoeriol, isosakuranetin, and nobiletin. In plants, O-methyltransferases (OMTs) participate in the methylation of a vast array of secondary metabolites, including flavonoids, phenylpropanoids, and alkaloids. To identify functional OMTs involved in the formation of methoxyflavonoids, orange (Citrus sinensis) OMT (CsOMT) genes were retrieved from the Citrus Genome Database. The phylogenetic relationships with functional OMTs suggested that three CsOMTs, CsOMT15, CsOMT16, and CsOMT30, are possible candidates for flavonoid OMTs (FOMTs). These CsOMTs were heterologously expressed in Escherichia coli, and their OMT activity was examined with flavonoid substrates. Of the examined CsOMTs, CsOMT16 catalyzed the regiospecific 3'-O-methylation of flavonoids to the respective 3'-methoxyflavonoids. A kinetic study demonstrated that CsOMT16 accepts diverse flavonoids as a substrate with a comparable preference. The flavonoids eriodictyol, luteolin, and quercetin were efficiently converted to homoeriodictyol, chrysoeriol, and isorhamnetin by CsOMT16-transformed E. coli cells, respectively. These findings suggest that CsOMT16 contributes to the methoxyflavonoid formation in orange and is applicable to the biotechnological production of 3'-methoxyflavonoids.

中文翻译：

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橙子区域特异性黄酮类 3'-O-甲基转移酶的生化表征

柑橘类植物含有多种甲氧基黄酮类化合物，包括金油醇、异樱素和川陈皮素。在植物中，O-甲基转移酶 (OMT) 参与大量次生代谢物的甲基化，包括类黄酮、苯丙素和生物碱。为了鉴定参与甲氧基黄酮类化合物形成的功能性 OMT，从柑橘基因组数据库中检索了橙（柑橘）OMT (CsOMT) 基因。与功能性 OMT 的系统发育关系表明，三个 CsOMT（CsOMT15、CsOMT16 和 CsOMT30）可能是类黄酮 OMT（FOMT）的候选者。这些 CsOMT 在大肠杆菌中异源表达，并用类黄酮底物检查了它们的 OMT 活性。在所检查的 CsOMT 中，CsOMT16 催化黄酮类化合物的区域特异性 3'-O-甲基化为相应的 3'-甲氧基黄酮类化合物。动力学研究表明，CsOMT16 以相当的偏好接受多种类黄酮作为底物。CsOMT16 转化的大肠杆菌细胞分别将类黄酮圣草酚、木犀草素和槲皮素有效转化为高圣草酚、金草醇和异鼠李素。这些发现表明，CsOMT16 有助于橙子中甲氧基黄酮类化合物的形成，并且适用于 3'-甲氧基黄酮类化合物的生物技术生产。