Enhanced production of antifungal lipopeptide iturin A by Bacillus amyloliquefaciens LL3 through metabolic engineering and culture conditions optimization
Enhanced production of antifungal lipopeptide iturin A by Bacillus amyloliquefaciens LL3 through metabolic engineering and culture conditions optimization
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通过代谢工程和培养条件优化，解淀粉芽孢杆菌 LL3 增强抗真菌脂肽伊枯草菌素 A 的生产
DOI：
10.1186/s12934-019-1121-1
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发表时间：
2019-04-10
影响因子：
6.4
通讯作者：
Yang, Chao
中科院分区：
文献类型：
--
作者：
Dang, Yulei;Zhao, Fengjie;Yang, Chao
背景：Iturins是一种主要由芽孢杆菌产生的抗生素类环状脂肽，因其具有溶血和抗真菌的特性，在生物医学和生物防治领域具有潜在的应用前景。本实验室先前分离到的解淀粉芽孢杆菌LL3具有完整的iturin a生物合成途径。然而，菌株LL3无法合成iturin A，可能是由于iturin A操纵子转录水平低所致。结果：在本研究中，通过在itu操纵子上游插入强组成启动子C2up，实现了iturin A生物合成基因的增强转录，生成了滴度为37.35 mg l-1的iturin A。液相色谱-质谱分析表明，菌株产生4个iturin A同源物，其分子离子峰在m/z 1044、1058、1072和1086处，分别为[C14+ 2H]2+、[C15+ 2H]2+、[C16+ 2H]2+和[C17+ 2H]2+。结果表明，iturin A提取物对几种常见植物病原菌具有较强的抑制活性。利用响应面法优化发酵条件，使iturin A的产率达到99.73 mg l-1。此外，通过过表达多效性调节因子DegQ， iturin A的产量增加到113.1 mg -1。结论：据我们所知，这是首次报道一株芽孢杆菌同时产生4种iturin A同源物（C14-C17）。此外，本研究表明，代谢工程与培养条件优化相结合可能是提高细菌次生代谢物产量的可行方法。
Background:Iturins, which belong to antibiotic cyclic lipopeptides mainly produced by Bacillus sp., have the potential for application in biomedicine and biocontrol because of their hemolytic and antifungal properties. Bacillus amyloliquefaciens LL3, isolated previously by our lab, possesses a complete iturin A biosynthetic pathway as shown by genomic analysis. Nevertheless, iturin A could not be synthesized by strain LL3, possibly resulting from low transcription level of the itu operon.Results:In this work, enhanced transcription of the iturin A biosynthetic genes was implemented by inserting a strong constitutive promoter C2up into upstream of the itu operon, leading to the production of iturin A with a titer of 37.35 mg l-1. Liquid chromatography-mass spectrometry analyses demonstrated that the strain produced four iturin A homologs with molecular ion peaks at m/z 1044, 1058, 1072 and 1086 corresponding to [C14+ 2H]2+, [C15+ 2H]2+, [C16+ 2H]2+and [C17+ 2H]2+. The iturin A extract exhibited strong inhibitory activity against several common plant pathogens. The yield of iturin A was improved to 99.73 mg l-1by the optimization of the fermentation conditions using a response surface methodology. Furthermore, the yield of iturin A was increased to 113.1 mg l-1by overexpression of a pleiotropic regulator DegQ.Conclusions:To our knowledge, this is the first report on simultaneous production of four iturin A homologs (C14-C17) by a Bacillus strain. In addition, this study suggests that metabolic engineering in combination with culture conditions optimization may be a feasible method for enhanced production of bacterial secondary metabolites.