Guanitrypmycin Biosynthetic Pathways Imply Cytochrome P450 Mediated Regio- and Stereospecific Guaninyl-Transfer Reactions
Guanitrypmycin Biosynthetic Pathways Imply Cytochrome P450 Mediated Regio- and Stereospecific Guaninyl-Transfer Reactions
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DOI：
10.1002/anie.201906891
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发表时间：
2019-07-05
影响因子：
16.6
通讯作者：
Li, Shu-Ming
中科院分区：
文献类型：
--
作者：
Liu, Jing;Xie, Xiulan;Li, Shu-Ming
挖掘包括链霉菌在内的微生物基因组揭示了大量生物合成基因簇的存在。揭示这种遗传潜力已被证明是发现新化合物的有用方法。在这里，我们报道了两个相似的含p450相关环二肽合成酶的基因簇在彩色链霉菌中的异源表达，并鉴定了8种罕见的新型天然产物，c3 -鸟嘌呤吲哚生物碱鸟嘌呤霉素。不同基因组合的表达证明，环二肽合成酶组装了环-l- trp -l- phe和环-l- trp -l- tyr，它们被环二肽氧化酶、细胞色素P450酶和n -甲基转移酶连续和区域特异性修饰。体内和体外实验结果证明，P450酶作为关键的生物催化剂，可催化色氨酸部分吲哚环上的区域特异性和立体特异性3 α -鸟嘌呤化。同位素交换实验为生物合成途径产物通过酮烯互变异构进行非酶外映提供了证据。这种途径后修饰在培养过程中进一步增加了胍霉素的结构多样性。
Mining microbial genomes including those of Streptomyces reveals the presence of a large number of biosynthetic gene clusters. Unraveling this genetic potential has proved to be a useful approach for novel compound discovery. Here, we report the heterologous expression of two similar P450-associated cyclodipeptide synthase-containing gene clusters in Streptomyces coelicolor and identification of eight rare and novel natural products, the C3-guaninyl indole alkaloids guanitrypmycins. Expression of different gene combinations proved that the cyclodipeptide synthases assemble cyclo-l-Trp-l-Phe and cyclo-l-Trp-l-Tyr, which are consecutively and regiospecifically modified by cyclodipeptide oxidases, cytochrome P450 enzymes, and N-methyltransferases. In vivo and in vitro results proved that the P450 enzymes function as key biocatalysts and catalyze the regio- and stereospecific 3 alpha-guaninylation at the indole ring of the tryptophanyl moiety. Isotope-exchange experiments provided evidence for the non-enzymatic epimerization of the biosynthetic pathway products via keto-enol tautomerism. This post-pathway modification during cultivation further increases the structural diversity of guanitrypmycins.