Elucidation of novel FPP-methyltransferase-terpene synthase gene clusters in Proteobacteria

传统的萜类化合物的生物合成是以C10-GPP、C15-FPP或C20-GGPP为底物的，但必须考虑范式的改变。FPP甲基转移酶的C-甲基化和环化与一个非经典的萜类合成酶的偶联代表了一种前所未有的反应机制，这是最近在褶皱沙雷氏菌4Rx13中独特的生物合成Sodorifen的反应机制。它说明了由于C16底物的利用，扩大萜类多样性的另一种途径。随后的Sodorifen合成酶对非规范的单环前Sodorifen焦磷酸盐的底物专一性意味着两种酶的共同进化。这种共同进化的初步证据也在产生新的双高萜类氯代菲的绿假单胞菌O6中发现。到目前为止，戊二烯基焦磷酸盐的C-甲基化和环化反应只在仔细研究各自生物合成途径来源的细菌中发现。我们推测，在蛋白质细菌而不是放线菌(如链霉菌)中，FPP的C-甲基化和环化是为了合成新的萜类化合物而建立的。在这里，我们将验证C17化合物的生物合成是否依赖于前面的两个甲基转移酶反应，然后是萜烯合成酶反应。此外，我们将检验FPP甲基转移酶在进化过程中是否(排他性/主要)在变形杆菌中表现出来。

The classical biosynthesis of terpenoids is based on the usage of C10-GPP, C15-FPP or C20-GGPP as substrates, but a paradigm change has to be considered. A coupling of C-methylation and cyclization by the FPP methyltransferase with a non-classical terpene synthase represents an unprecedented reaction mechanism which was recently shown for the unique biosynthesis of sodorifen in Serratia plymuthica 4Rx13. It illustrates an alternative avenue to enlarge terpene diversity due to the utilization of a C16 substrate. Substrate specificity of the succeeding sodorifen synthase for the non-canonical, monocyclic pre-sodorifen pyrosphosphate implies the coevolution of both enzymes. Preliminary additional evidence of this coevolution is also found in Pseudomonas chlororaphis O6 which produces the novel bis-homoterpene chlororaphen. So far, C-methylations and cyclizations of prenyl pyrophosphates have been exclusively found in bacteria scrutinizing the origin of respective biosynthetic pathways. We hypothesize that in proteobacteria rather than in actinobacteria (e.g. Streptomyces species) C-methylation and cyclization of FPP was established for the biosynthesis of novel terpenoids. Here we will verify whether the biosynthesis the of C17 compound chlororaphen depends on two preceding methyltransferase reactions which are followed by a terpene synthase reaction. Moreover, we will test the hypothesis whether FPP methyltransferases were (exclusively/dominantly) manifested in proteobacteria during evolution.