Kazan Institute of Biochemistry and Biophysics, Russian Academy of Sciences, P.O. Box 30, Kazan, 420111, Russia.
Kazan Institute of Biochemistry and Biophysics, Russian Academy of Sciences, P.O. Box 30, Kazan, 420111, Russia.
Phytochemistry. 2018 Dec;156:73-82. doi: 10.1016/j.phytochem.2018.08.010. Epub 2018 Sep 6.
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 H-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.
模式植物石松（Selaginella moellendorffii Hieron）的 CYP74M2 基因被克隆，研究了相应重组蛋白的催化特性。重组 CYP74M2 蛋白对亚油酸和 a-亚麻酸的 13-过氧化物（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 不同，CYP74M2 表现为专用环氧化物合酶（EAS）。例如，13-HPOD 转化为三种差向异构的环氧醇 1-3（形成比例约为 4:2:1），即（11R,12S,13S）、（11R,12R,13S）和（11S,12S,13S）（9Z）-11-羟基-12,13-环氧-9-十八烯酸。除了这些产物，还形成了少数环氧乙烯基醇，如（10E）-11-羟基-12,13-环氧-9-十八烯酸。CYP74M2 转化 13-HPOT 生成 11-羟基-12,13-环氧-9,15-十八碳二烯酸的两种立体异构体。通过 HPLC 纯化各个氧脂素，最终通过其 NMR 数据，包括 H-NMR、2D-COSY、HSQC 和 HMBC 进行鉴定。因此，CYP74M2 是专用环氧化物合酶。据我们所知，目前尚未在高等植物中检测到这种类型的酶。
