The CYP74M2 gene of a model plant, the spikemoss Selaginella moellendorffii Hieron, was cloned and the catalytic properties of corresponding recombinant protein were studied. The recombinant CYP74M2 protein was active towards 13-hydroperoxides of linoleic and a-linolenic acids (13-HPOD and 13-HPOT, respectively). In contrast to previously studied CYP74M1 and CYP74M3, which possessed the divinyl ether synthase activity, CYP74M2 behaved as a dedicated epoxyalcohol synthase (EAS). For instance, the 13-HPOD was converted to three epimeric oxiranyl carbinols 1-3 (formed at a ratio ca. 4:2:1), namely the (11R,12S,13S), (11R,12R, 13S), and (11S,12S,13S) epimers of (9Z)-11-hydroxy-12,13-epoxy-9-octadecenoic acid. Besides these products, a minority of oxiranyl vinyl carbinols like (10E)-11-hydroxy-12,13-epoxy-9-octadecenoic acid was formed. The 13-HPOT conversion by CYP74M2 afforded two stereoisomers of 11-hydroxy-12,13-epoxy-9,15-octadecadienoic acid. Individual oxylipins were purified by HPLC and finally identified by their NMR data, including the 1H-NMR, 2D-COSY, HSQC, and HMBC. Thus, the CYP74M2 is the dedicated epoxyalcohol synthase. To our knowledge, no enzymes of this type have been detected in higher plants yet.

中文翻译：

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第一个高等植物环氧醇合酶的检测：刺苔卷柏(Selaginella moellendorffii) CYP74M2酶的分子克隆和表征

克隆了模式植物Selaginella moellendorffii Hieron的CYP74M2基因，并研究了相应重组蛋白的催化特性。重组 CYP74M2 蛋白对亚油酸和α-亚麻酸的 13-氢过氧化物（分别为 13-HPOD 和 13-HPOT）具有活性。与先前研究的具有二乙烯基醚合酶活性的 CYP74M1 和 CYP74M3 相比，CYP74M2 表现为专用的环氧醇合酶 (EAS)。例如，13-HPOD 被转化为三个差向异构环氧乙烷基甲醇 1-3（以大约 4:2:1 的比例形成），即 (11R,12S,13S)、(11R,12R,13S)，和(9Z)-11-羟基-12,13-环氧-9-十八碳烯酸的(11S,12S,13S)差向异构体。除了这些产物外，还形成了少数环氧乙烷基乙烯基甲醇，如 (10E)-11-羟基-12,13-环氧-9-十八碳烯酸。CYP74M2 的 13-HPOT 转化提供了 11-羟基-12,13-环氧-9,15-十八碳二烯酸的两种立体异构体。单个 oxylipins 通过 HPLC 纯化，最后通过它们的 NMR 数据进行鉴定，包括 1H-NMR、2D-COSY、HSQC 和 HMBC。因此，CYP74M2 是专用的环氧醇合酶。据我们所知，尚未在高等植物中检测到此类酶。