Pyocin S5 Import into Pseudomonas aeruginosa Reveals a Generic Mode of Bacteriocin Transport
Pyocin S5 Import into Pseudomonas aeruginosa Reveals a Generic Mode of Bacteriocin Transport
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DOI：
10.1128/mbio.03230-19
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发表时间：
2020-03-01
期刊：
影响因子：
6.4
通讯作者：
Kleanthous, Colin
中科院分区：
文献类型：
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作者：
Behrens, Hannah M.;Lowe, Edward D.;Kleanthous, Colin
绿脓菌素S5（PyoS 5）是一种有效的蛋白质细菌素，在动物感染模型中根除人类病原体铜绿假单胞菌，但其输入机制知之甚少。在这里，使用晶体学，生物物理和生物化学分析，以及活细胞成像，我们定义了PyoS 5的进入过程，并揭示了与其他细菌素的运输机制的联系。除了其C-末端成孔结构域之外，延长的PyoS 5还包含两个新的串联重复的扭结的3-螺旋束结构域，基于结构的比对将其鉴定为其他绿脓菌素中的关键输入结构域。中心结构域结合脂质结合的常见多糖抗原，允许绿脓菌素在细胞表面上积累。N-末端结构域结合铁绿脓菌螯铁蛋白转运蛋白FptA，而其相关的无序区域结合内膜蛋白TonB 1，它们一起驱动细菌素穿过外膜的输入。最后，我们确定了致敏大肠杆菌对PyoS 5，以及其他绿脓菌素的最低要求，并建议一个通用的途径可能支持进口的所有TonB依赖性细菌素的外膜革兰氏阴性bacterios.IMPORTANCE细菌素是有毒的多肽细菌杀死他们的竞争对手，使他们有趣的潜在抗生素。在这里，我们揭示了细菌素摄取途径的意料之外的共性，通过分子和细胞解剖的进口途径的成孔细菌素绿脓菌素S5（PyoS 5），其目标是铜绿假单胞菌。除了其C-末端孔形成结构域之外，PyoS 5还由两个串联重复的螺旋结构域组成，这也是我们在其他绿脓菌素中鉴定到的。功能分析表明，它们在进口过程中发挥着不同的作用。一个识别从表面投射的保守糖，而另一个识别特定的外膜铁载体转运蛋白FptA，在PyoS 5的情况下。通过对大肠杆菌细胞的工程改造，我们表明绿脓菌素可以很容易地被重新利用来杀死其他物种。这表明外膜易位步骤的基本规则可能适用于许多革兰氏阴性细菌的细菌素。
Pyocin S5 (PyoS5) is a potent protein bacteriocin that eradicates the human pathogen Pseudomonas aeruginosa in animal infection models, but its import mechanism is poorly understood. Here, using crystallography, biophysical and biochemical analyses, and live-cell imaging, we define the entry process of PyoS5 and reveal links to the transport mechanisms of other bacteriocins. In addition to its C-terminal pore-forming domain, elongated PyoS5 comprises two novel tandemly repeated kinked 3-helix bundle domains that structure-based alignments identify as key import domains in other pyocins. The central domain binds the lipid-bound common polysaccharide antigen, allowing the pyocin to accumulate on the cell surface. The N-terminal domain binds the ferric pyochelin transporter FptA while its associated disordered region binds the inner membrane protein TonB1, which together drive import of the bacteriocin across the outer membrane. Finally, we identify the minimal requirements for sensitizing Escherichia coli toward PyoS5, as well as other pyocins, and suggest that a generic pathway likely underpins the import of all TonB-dependent bacteriocins across the outer membrane of Gram-negative bacteria.IMPORTANCE Bacteriocins are toxic polypeptides made by bacteria to kill their competitors, making them interesting as potential antibiotics. Here, we reveal unsuspected commonalities in bacteriocin uptake pathways, through molecular and cellular dissection of the import pathway for the pore-forming bacteriocin pyocin S5 (PyoS5), which targets Pseudomonas aeruginosa. In addition to its C-terminal poreforming domain, PyoS5 is composed of two tandemly repeated helical domains that we also identify in other pyocins. Functional analyses demonstrate that they have distinct roles in the import process. One recognizes conserved sugars projected from the surface, while the other recognizes a specific outer membrane siderophore transporter, FptA, in the case of PyoS5. Through engineering of Escherichia coli cells, we show that pyocins can be readily repurposed to kill other species. This suggests basic ground rules for the outer membrane translocation step that likely apply to many bacteriocins targeting Gram-negative bacteria.