A Peptidomimetic Antibiotic Interacts with the Periplasmic Domain of LptD from Pseudomonas aeruginosa
A Peptidomimetic Antibiotic Interacts with the Periplasmic Domain of LptD from Pseudomonas aeruginosa
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DOI：
10.1021/acschembio.7b00822
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发表时间：
2018-03-01
影响因子：
4
通讯作者：
Robinson, John A.
中科院分区：
文献类型：
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作者：
Andolina, Gloria;Bencze, Laszlo-Csaba;Robinson, John A.
革兰氏阴性菌的外膜（OM）是一个不对称的双层，主要是脂多糖（LPS）分子在外小叶。在OM生物发生过程中，新的LPS分子通过七种LPS转运蛋白（LptA-G）从其在内膜上的组装位点转运到OM。在整合的β-桶OM蛋白LptD和脂蛋白LptE之间形成的复合物负责将LPS从OM的周质侧转运到其在细胞表面上的最终位置。由于其在许多革兰氏阴性菌中的重要功能，LPS转运途径是开发新抗生素的有趣靶点。最近发现了一个家族的大环肽模拟物，其靶向LptD并特异性地抑制假单胞菌中的LPS转运。相关分子Murepavadin正在临床开发中，用于治疗由铜绿假单胞菌引起的危及生命的感染。为了表征这些抗生素与来自铜绿假单胞菌的LptD的相互作用，我们通过与光标记探针交联来表征结合位点。我们使用了一种基于质谱的无假设蛋白质组学方法来提供证据，证明抗生素与LptD的周质片段交联，该周质片段含有假单胞菌特有的β-淀粉样结构域和N-末端插入结构域。预期抗生素与周质段的结合可阻断LPS转运，这与这些抗生素的拟定作用模式和观察到的特异性一致。这些见解可能对发现靶向其他革兰氏阴性菌中LPS转运途径的新抗生素有价值。
The outer membrane (OM) in Gram-negative bacteria is an asymmetric bilayer with mostly lipopolysaccharide (LPS) molecules in the outer leaflet. During OM biogenesis, new LPS molecules are transported from their site of assembly on the inner membrane to the OM by seven LPS transport proteins (LptA-G). The complex formed between the integral beta-barrel OM protein LptD and the lipoprotein LptE is responsible for transporting LPS from the periplasmic side of the OM to its final location on the cell surface. Because of its essential function in many Gram-negative bacteria, the LPS transport pathway is an interesting target for the development of new antibiotics. A family of macrocyclic peptidomimetics was discovered recently that target LptD and inhibit LPS transport specifically in Pseudomonas spp. The related molecule Murepavadin is in clinical development for the treatment of life-threatening infections caused by P. aeruginosa. To characterize the interaction of these antibiotics with LptD from P. aeruginosa, we characterized the binding site by cross-linking to a photolabeling probe. We used a hypothesis-free mass spectrometry-based proteomic approach to provide evidence that the antibiotic cross-links to the periplasmic segment of LptD, containing a beta-jellyroll domain and an N-terminal insert domain characteristic of Pseudomonas spp. Binding of the antibiotic to the periplasmic segment is expected to block LPS transport, consistent with the proposed mode of action and observed specificity of these antibiotics. These insights may prove valuable for the discovery of new antibiotics targeting the LPS transport pathway in other Gram-negative bacteria.