Laboratory for Bioanalytical Chemistry, Institute of Chemistry , University of Neuchatel , Avenue de Bellevaux 51 , CH-2000 Neuchâtel , Switzerland.
Department of Bioorganic Chemistry , Max Planck Institute for Chemical Ecology , Hans-Knoell-Straße 8 , D-07745 Jena , Germany.
J Am Chem Soc. 2018 Sep 19;140(37):11855-11862. doi: 10.1021/jacs.8b08510. Epub 2018 Sep 4.
The rhizobacterium Serratia plymuthica 4Rx13 releases a unique polymethylated hydrocarbon (CH) with a bicyclo[3.2.1]octadiene skeleton called sodorifen. Sodorifen production depends on a gene cluster carrying a C-methyltransferase and a terpene cyclase along with two enzymes of the 2- C-methyl-d-erythritol 4-phosphate (MEP) pathway of isoprenoid biosynthesis. Comparative analysis of wild-type and mutant volatile organic compound profiles revealed a C-methyltransferase-dependent C alcohol called pre-sodorifen, the production of which is upregulated in the terpene cyclase mutant. The monocyclic structure of this putative intermediate in sodorifen biosynthesis was identified by NMR spectroscopy. In vitro assays with the heterologously expressed S. plymuthica C-methyltransferase and terpene cyclase demonstrated that these enzymes act sequentially to convert farnesyl pyrophosphate (FPP) into sodorifen via a pre-sodorifen pyrophosphate intermediate, indicating that the S-adenosyl methionine (SAM)-dependent C-methyltransferase from S. plymuthica exhibits unprecedented cyclase activity. In vivo incorporation experiments with C-labeled succinate, l-alanine, and l-methionine confirmed a MEP pathway to FPP via the canonical glyceraldehyde-3-phosphate and pyruvate, as well as its SAM-dependent methylation in pre-sodorifen and sodorifen biosynthesis. C{H} NMR spectroscopy facilitated the localization of C labels and provided detailed insights into the biosynthetic pathway from FPP via pre-sodorifen pyrophosphate to sodorifen.
植物根际促生菌粘质沙雷氏菌 4Rx13 会释放一种独特的多甲基化碳氢化合物（CH），骨架为双环[3.2.1]辛二烯，名为 sodorifen。sodorifen 的产生依赖于一个基因簇，该基因簇携带有 C-甲基转移酶和萜烯环化酶，以及异戊烯基生物合成的 2-C-甲基-D-赤藓糖醇 4-磷酸（MEP）途径的两种酶。野生型和突变体挥发性有机化合物谱的比较分析显示，有一种依赖 C-甲基转移酶的 C-醇，称为 pre-sodorifen，萜烯环化酶突变体中其产量上调。该化合物在 sodorifen 生物合成中的中间产物的单环结构通过 NMR 光谱确定。用异源表达的粘质沙雷氏菌 C-甲基转移酶和萜烯环化酶进行的体外测定表明，这些酶依次作用，通过 pre-sodorifen 焦磷酸中间产物将法呢基焦磷酸（FPP）转化为 sodorifen，表明粘质沙雷氏菌的 S-腺苷甲硫氨酸（SAM）依赖性 C-甲基转移酶表现出前所未有的环化酶活性。用 C 标记琥珀酸、l-丙氨酸和 l-蛋氨酸进行的体内掺入实验证实了通过经典的甘油醛-3-磷酸和丙酮酸以及其在 pre-sodorifen 和 sodorifen 生物合成中的 SAM 依赖性甲基化作用将 MEP 途径转化为 FPP。C{13}H}NMR 光谱有助于定位 C 标记，并提供了详细的见解，了解了从 FPP 通过 pre-sodorifen 焦磷酸到 sodorifen 的生物合成途径。
