Deletion of COM donor and acceptor domains and the interaction between modules in bacillomycin D produced by Bacillus amyloliquefaciens.
Deletion of COM donor and acceptor domains and the interaction between modules in bacillomycin D produced by Bacillus amyloliquefaciens.
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DOI：
10.1016/j.synbio.2022.05.007
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发表时间：
2022-09
影响因子：
4.8
通讯作者：
Bie, Xiaomei
中科院分区：
文献类型：
--
作者：
Lv, Ziyan;Ma, Wenjie;Zhang, Ping;Lu, Zhaoxin;Zhou, Libang;Meng, Fanqiang;Wang, Zuwei;Bie, Xiaomei
Bacillomycin D 是由解淀粉芽孢杆菌 fmbJ 产生的环状脂肽。目前，还没有相关报道描述芽孢杆菌D的组合生物合成。由于通信介导（COM）结构域具有强大的生物合成潜力，其NRPS亚基之间的串扰已得到一定程度的研究，但COM结构域在模块之间的相互作用却很少报道。因此，在本研究中，我们通过删除模块之间的COM供体和受体结构域进行了杆菌霉素D的组合生物合成，并阐明了NRPS模块之间的相互作用。结果表明，模块2和模块3之间供体结构域的缺失并没有影响上游模块的催化作用，但阻止了下游模块催化脂肽产物的延伸，最终产生能够形成序列为β-NH2FA-Asn-Tyr的线性二肽的突变复合物。然而，在模块 3 和 4 以及模块 6 和 7 之间缺乏供体结构域的工程杂交芽孢杆菌素 D NRPS 可以形成多个装配线，生产芽孢杆菌素 D 及其类似物（线性三肽、环状六肽和线性六肽）。此外，所有受体域缺失菌株均不能产生芽孢杆菌素D，仅产生模块中断产生的截短肽（除了模块3和4之间的受体域缺失菌株也产生环六肽）。总之，模块间供体结构域的删除导致了更灵活的混合生物合成系统，用于生产多种肽产品；与只能产生截短肽的亚基间供体结构域缺失菌株相比，前者具有更强的生物合成能力。同时，模块之间的受体结构域是杆菌素D合成酶内模块间相互作用和有效通讯的重要组成部分。
Bacillomycin D is a cyclic lipopeptide produced by Bacillus amyloliquefaciens fmbJ. At present, no relevant report has described the combinatorial biosynthesis of bacillomycin D. Due to the strong biosynthetic potential of the communication-mediating (COM) domains, its crosstalk between NRPS subunits has been studied to some extent, but the interaction of COM domain between modules is rarely reported. Therefore, in this study, we conducted the combinatorial biosynthesis of bacillomycin D through the deletion of the COM donor and acceptor domains between the modules and elucidated the interaction between the NRPS modules. The results showed that the deletion of the donor domain between modules 2 and 3 did not affect catalysis by upstream modules, but prevented downstream modules from catalysing the extension of the lipopeptide product, ultimately resulting in mutant complexes that could form linear dipeptides with the sequence β-NH2FA-Asn-Tyr. However, the engineered hybrid bacillomycin D NRPSs lacking the donor domains between modules 3 and 4 and modules 6 and 7 could form multiple assembly lines that produced bacillomycin D and its analogs (linear tripeptides, cyclic hexapeptides and linear hexapeptides). In addition, all the acceptor domain deletion strains failed to produce bacillomycin D, only truncated peptides produced by module interruption (except for the acceptor domain deletion strains between modules 3 and 4, which also produced cyclic hexapeptides). In conclusion, deletion of the inter-module donor domains led to a more flexible hybrid biosynthetic system for the production of diverse peptide products; compared with the inter-subunit donor domain deletion strains that could only produce truncated peptides, the former had a greater biosynthetic capacity. Meanwhile, the acceptor domains between modules were an important part of module-module interactions and efficient communication within bacillomycin D synthetase.
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