Two Functional Fatty Acyl Coenzyme A Ligases Affect Free Fatty Acid Metabolism To Block Biosynthesis of an Antifungal Antibiotic in Lysobacter enzymogenes
Two Functional Fatty Acyl Coenzyme A Ligases Affect Free Fatty Acid Metabolism To Block Biosynthesis of an Antifungal Antibiotic in Lysobacter enzymogenes
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两种功能性脂肪酰辅酶 A 连接影响游离脂肪酸代谢，阻断溶杆菌酶基因中抗真菌抗生素的生物合成
DOI：
10.1128/aem.00309-20
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发表时间：
2020-05-01
影响因子：
4.4
通讯作者：
Liu, Fengquan
中科院分区：
文献类型：
--
作者：
Li, Kaihuai;Hou, Rongxian;Liu, Fengquan
在产酶溶杆菌OH 11中，RpfB 1和RpfB 2被预测编码酰基辅酶A（CoA）连接酶。RpfB 1位于Rpf基因簇中。有趣的是，我们在这个典型基因簇之外发现了一个RpfB 1同源物（RpfB 2），但对其功能或机制一无所知。在这里，我们报告，rpfB 1和rpfB 2可以在功能上取代大肠杆菌fadD突变体JW 1794中的EcFadD。RpfB在体外激活长链脂肪酸（n-C-16：0和n-C-18（：0））以获得相应的脂肪酰辅酶A连接酶（FCL）活性，而Glu-361在RpfB 1和RpfB 2的催化机制中起关键作用。rpf 81和rpf 82的缺失导致热稳定抗真菌因子（HSAF）的产生显著增加，rpf 81或rpfB 2的过表达完全抑制HSAF的产生。rpfB 1和rpfB 2的缺失导致L.酶基因扩散信号因子3（LeDSF 3）的合成。酶基因总之，我们的研究结果表明，细胞内游离脂肪酸水平的变化显着改变HSAF的生产。我们的报告表明，细胞内游离脂肪酸是HSAF生产所必需的，RpfB通过FCL活性影响HSAF生产。全局转录调控因子Clp直接调控rpfB 1和rpfB 2的表达。这些结果揭示了RpfB在L.重要性了解热稳定抗真菌因子（HSAF）的生物合成和调控机制可以提高产酶溶杆菌的产量。在这里，我们报告RpfB 1和RpfB 2编码酰基辅酶A（CoA）连接酶。我们的研究表明，RpfB 1和RpfB 2通过脂肪酰辅酶A连接酶（FCL）活性影响游离脂肪酸代谢，从而减少HSAF合成的底物，从而阻断L.酶基因此外，这些发现揭示了脂肪酰辅酶A连接酶RpfB 1和RpfB 2在L.酶基因重要的是，这项工作的新奇是发现RpfB 2位于Rpf基因簇之外，并在HSAF生产中起关键作用，这在其他扩散信号因子（DSF）/Rpf生产细菌中尚未报道。
In Lysobacter enzymogenes OH11, RpfB1 and RpfB2 were predicted to encode acyl coenzyme A (CoA) ligases. RpfB1 is located in the Rpf gene cluster. Interestingly, we found an RpfB1 homolog (RpfB2) outside this canonical gene cluster, and nothing is known about its functionality or mechanism. Here, we report that rpfB1 and rpfB2 can functionally replace EcFadD in the Escherichia coil fadD mutant JW1794. RpfB activates long-chain fatty acids (n-C-16:0 and n-C-18(:0)) for the corresponding fatty acyl-CoA ligase (FCL) activity in vitro, and Glu-361 plays critical roles in the catalytic mechanism of RpfB1 and RpfB2. Deletion of rpf81 and rpf82 resulted in significantly increased heat-stable antifungal factor (HSAF) production, and overexpression of rpf81 or rpfB2 completely suppressed HSAF production. Deletion of rpfB1 and rpfB2 resulted in increased L. enzymogenes diffusible signaling factor 3 (LeDSF3) synthesis in L. enzymogenes. Overall, our results showed that changes in intracellular free fatty acid levels significantly altered HSAF production. Our report shows that intracellular free fatty acids are required for HSAF production and that RpfB affects HSAF production via FCL activity. The global transcriptional regulator Clp directly regulated the expression of rpfB1 and rpfB2. In conclusion, these findings reveal new roles of RpfB in antibiotic biosynthesis in L. enzymogenes.IMPORTANCE Understanding the biosynthetic and regulatory mechanisms of heat-stable antifungal factor (HSAF) could improve the yield in Lysobacter enzymogenes. Here, we report that RpfB1 and RpfB2 encode acyl coenzyme A (CoA) ligases. Our research shows that RpfB1 and RpfB2 affect free fatty acid metabolism via fatty acyl-CoA ligase (FCL) activity to reduce the substrate for HSAF synthesis and, thereby, block HSAF production in L. enzymogenes. Furthermore, these findings reveal new roles for the fatty acyl-CoA ligases RpfB1 and RpfB2 in antibiotic biosynthesis in L. enzymogenes. Importantly, the novelty of this work is the finding that RpfB2 lies outside the Rpf gene cluster and plays a key role in HSAF production, which has not been reported in other diffusible signaling factor (DSF)/Rpf-producing bacteria.