Secondary metabolites overproduction through transcriptional gene cluster refactoring
Secondary metabolites overproduction through transcriptional gene cluster refactoring
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DOI：
10.1016/j.ymben.2018.09.010
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发表时间：
2018-09-01
影响因子：
8.4
通讯作者：
Luzhetskyy, Andriy
中科院分区：
文献类型：
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作者：
Horbal, Liliya;Marques, Filipe;Luzhetskyy, Andriy
我们提出了一个随机的合理的方法，使建设生产过剩的菌株在两个步骤。该方法首先涉及创建感兴趣的簇的文库，其中天然启动子被随机产生的组成型合成启动子取代，然后在适当的宿主菌株中表达该文库。这种策略快速、易于使用，解释了簇的结构，并将基因簇的表达与复杂的天然调控网络完全脱钩。该策略被应用于提高大环肽botromycin的生产，botromycin对多重耐药细菌具有抗菌活性，是一类新的抗菌药物的蓝图。我们成功地优化了操纵子中基因的表达，并产生了几种波曲霉素基因簇的变体，这些变体提供了比天然的波曲霉素高5-50倍的滴度，从而鉴定了几种以前没有描述过的新的波曲霉素衍生物。此外，由于更高的博曲霉素产率，通过基因簇的生物合成工程进行博曲霉素衍生化。上述特征使得这种通用策略成为放线菌中过量生产已知次级代谢产物和激活沉默次级代谢产物的有前途的工具。
We present a random rational approach enabling the construction of overproducing strains in two steps. The approach first involves creating a library of clusters of interest, in which native promoters are substituted with randomly generated constitutive synthetic promoters, and then expressing this library in an appropriate host strain. This strategy is fast, easy to use, accounts for the architecture of a cluster and completely decouples the expression of a gene cluster from complex native regulatory networks. The strategy was applied to improve the production of a macrocyclic peptide, bottromycin, which possesses antibacterial activity against multidrug-resistant bacteria and is a blueprint for a new class of antibacterials. We successfully optimized the expression of genes in operons and created several variants of the bottromycin gene cluster that provide 5-50 fold higher titres of bottromycin than the natural one, thus resulting in the identification of several new bottromycin derivatives not previously described. Moreover, due to the higher bottromycin yield, bottromycin derivatization was performed via the biosynthetic engineering of the gene cluster. The abovementioned features make this generic strategy a promising tool for the overproduction of known secondary metabolites and the activation of silent secondary metabolites in Actinobacteria.