Directed natural product biosynthesis gene cluster capture and expression in the model bacterium Bacillus subtilis.
Directed natural product biosynthesis gene cluster capture and expression in the model bacterium Bacillus subtilis.
复制标题
定向的天然产物生物合成基因簇捕获和模型细菌枯草菌中的表达。
DOI：
10.1038/srep09383
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发表时间：
2015-03-24
影响因子：
4.6
通讯作者：
Qian PY
中科院分区：
文献类型：
--
作者：
Li Y;Li Z;Yamanaka K;Xu Y;Zhang W;Vlamakis H;Kolter R;Moore BS;Qian PY
芽孢杆菌是普遍存在的低G+C环境革兰氏阳性细菌，其产生各种各样的特化小分子。虽然它们的天然产物生物合成潜力很高，但支持芽孢杆菌宿主中大生物合成基因簇的异源表达的稳健分子工具是罕见的。本文中，我们采用酵母中的转化相关重组（TAR）来设计用于枯草芽孢杆菌中抗生素生产的单个基因组捕获和表达载体。在用表面活性素验证这种直接克隆“即插即用”方法后，我们从海洋分离物枯草芽孢杆菌1779中遗传地询问amicoumacin生物合成基因簇。它的异源表达使我们能够探索一个不寻常的成熟过程，涉及N-酰基-天冬酰胺前药中间体amicoumacins，这是由天冬酰胺特异性肽酶水解成活性成分amicoumacin A。这项工作代表了第一个直接克隆的基础上异源表达的天然产物在模式生物B。枯草杆菌和铺平了道路，未来的基因组挖掘工作在这一属的发展。
Bacilli are ubiquitous low G+C environmental Gram-positive bacteria that produce a wide assortment of specialized small molecules. Although their natural product biosynthetic potential is high, robust molecular tools to support the heterologous expression of large biosynthetic gene clusters in Bacillus hosts are rare. Herein we adapt transformation-associated recombination (TAR) in yeast to design a single genomic capture and expression vector for antibiotic production in Bacillus subtilis. After validating this direct cloning “plug-and-play” approach with surfactin, we genetically interrogated amicoumacin biosynthetic gene cluster from the marine isolate Bacillus subtilis 1779. Its heterologous expression allowed us to explore an unusual maturation process involving the N-acyl-asparagine pro-drug intermediates preamicoumacins, which are hydrolyzed by the asparagine-specific peptidase into the active component amicoumacin A. This work represents the first direct cloning based heterologous expression of natural products in the model organism B. subtilis and paves the way to the development of future genome mining efforts in this genus.