Characterization of the Polymyxin D Synthetase Biosynthetic Cluster and Product Profile of Paenibacillus polymyxa ATCC 10401
Characterization of the Polymyxin D Synthetase Biosynthetic Cluster and Product Profile of Paenibacillus polymyxa ATCC 10401
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DOI：
10.1021/acs.jnatprod.6b00807
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发表时间：
2017-05-01
影响因子：
5.1
通讯作者：
Velkov, Tony
中科院分区：
文献类型：
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作者：
Galea, Charles A.;Han, Meiling;Velkov, Tony
多粘菌素耐药细菌的日益流行刺激了对改进多粘菌素脂肽的研究。本文描述了多粘类芽孢杆菌ATCC 10401多粘菌素D非核糖体肽合成酶的序列和产物。多粘菌素D合成酶基因簇由5个基因组成，编码ABC转运蛋白(pmxC和pmxD)和负责多粘菌素D生物合成的酶(pmxA、pmxB和pmxE)。与多粘菌素B和E不同，多粘菌素D在第3位含有D-丝氨酸，而不是L-α-二氨基丁酸，在第7位有L-苏氨酸，而不是L-亮氨酸。PmxE的模块3含有一个辅助的异构化结构域，可催化L-丝氨酸向D-型的转化。结构模拟表明，PmxE的模块3和PmxA的模块6和7的腺化结构域可以与氨基酸结合，侧链比它们的首选底物更大。在培养基中添加单独的氨基酸不仅影响多粘菌素D-1和D-2的产生，还导致多粘菌素D第3、6和7位不同氨基酸的掺入。有趣的是，非天然多粘菌素类似物对一组革兰氏阴性临床分离株没有显示出抗菌活性，而天然多粘菌素D-1和D-2在小鼠血液感染模型中表现出良好的体外抗菌活性，对肺炎克雷伯菌和鲍曼不动杆菌有良好的抗菌活性。这些结果证明了这些不寻常的D-Ser(3)多粘菌素具有良好的抗菌活性，并强调了通过操纵多粘菌素非核糖体生物合成机制在多粘菌素D的第3、6和7位引入交替氨基酸的可能性。
The increasing prevalence of polymyxin-resistant bacteria has stimulated the search for improved polymyxin lipopeptides. Here we describe the sequence and product profile for polymyxin D nonribosomal peptide synthetase from Paenibacillus polymyxa ATCC 10401. The polymyxin D synthase gene cluster comprised five genes that encoded ABC transporters (pmxC and pmxD) and enzymes responsible for the biosynthesis of polymyxin D (pmxA, pmxB, and pmxE). Unlike polymyxins B and E, polymyxin D contains D-Ser at position 3 as opposed to L-alpha,gamma-diaminobutyric acid and has an L-Thr at position 7 rather than L-Leu. Module 3 of pmxE harbored an auxiliary epimerization domain that catalyzes the conversion of L-Ser to the D-form. Structural modeling suggested that the adenylation domains of module 3 in PmxE and modules 6 and 7 in PmxA could bind amino acids with larger side chains than their preferred substrate. Feeding individual amino acids into the culture media not only affected production of polymyxins D-1 and D-2 but also led to the incorporation of different amino acids at positions 3, 6, and 7 of polymyxin D. Interestingly, the unnatural polymyxin analogues did not show antibiotic activity against a panel of Gram-negative clinical isolates, while the natural polymyxins D-1 and D-2 exhibited excellent in vitro antibacterial activity and were efficacious against Klebsiella pneumoniae and Acinetobacter baumannii in a mouse blood infection model. The results demonstrate the excellent antibacterial activity of these unusual D-Ser(3) polymxyins and underscore the possibility of incorporating alternate amino acids at positions 3, 6, and 7 of polymyxin D via manipulation of the polymyxin nonribosomal biosynthetic machinery.