Gram-negative outer and inner membrane models: Insertion of cyclic cationic lipopeptides
Gram-negative outer and inner membrane models: Insertion of cyclic cationic lipopeptides
复制标题
DOI：
10.1021/jp064757
复制
发表时间：
2007-01-25
影响因子：
3.3
通讯作者：
Cajal, Yolanda
中科院分区：
文献类型：
--
作者：
Clausell, Adria;Garcia-Subirats, Maria;Cajal, Yolanda
大多数革兰氏阴性菌对多粘菌素B(PxB)敏感，对这种阳离子脂肽产生耐药性的情况非常罕见。PxB的作用机制涉及与细菌的外膜(OM)和内膜(IM)的相互作用。为了根据PxB的结构和改善的治疗指标设计新的抗生素，确定PxB的关键活性特征是必不可少的。我们使用了一种方法，分别用单层脂多糖或阴离子1-palmitoyl-2-oleoylglycero-sn-3-phosphoglycerol模拟革兰氏阴性菌OM和IM的外层，并结合渗透试验、压力/面积曲线分析和布鲁斯特角显微镜来监测表面形态的变化。合成类似物SP-B保持了天然化合物的基本结构特征，并以类似的方式与OM和IM相互作用。类似物SP-C将序列[D-Phe(6)-Leu(7)]突变为[D-Phe(6)-dab(7)]，表明该疏水结构域参与了内毒素的结合。用SP-DAP类似物证明了正电荷的显著作用，其中L-α，γ-二氨基丁酸残基DAB(1)和DAB(8)被L-α，伽马二氨基丙酸(DAP)取代，导致在LPS和PG单层中的插入度降低。多粘菌素B九肽(PxB-NP)证明了N-端酰化链的重要性。与PxB、Sp-B或Sp-C相比，PxB-NP对内毒素的亲和力较低，但它不能插入PG单层，尽管它表面上与阴离子膜结合。由于PxB微生物的杀灭似乎是由OM-IM磷脂交换引起的渗透不稳定性所介导的，因此利用磷脂单层囊泡研究了不同多肽诱导膜-膜脂交换的能力。结果表明，阳离子两亲性决定了多肽的活性。
Most Gram-negative bacteria are susceptible to polymyxin B (PxB), and development of resistance to this cationic lipopeptide is very rare. PxB mechanism of action involves interaction with both the outer membrane (OM) and the inner membrane (IM) of bacteria. For the design of new antibiotics based on the structure of PxB and with improved therapeutic indexes, it is essential to establish the key features of PxB that are important for activity. We have used an approach based on mimicking the outer layers of the OM and the IM of Gram-negative bacteria using monolayers of lipopolysaccharide (LPS) or anionic 1-palmitoyl-2-oleoylglycero-sn-3-phosphoglycerol (POPG), respectively, and using a combination of penetration assay, analysis of pressure/area curves, and Brewster angle microscopy to monitor surface morphology changes. Synthetic analogue sp-B maintains the basic structural characteristics of the natural compound and interacts with the OM and the IM in a similar way. Analogue sp-C, with a mutation of the sequence [D-Phe(6)-Leu(7)] into [D-Phe(6)-Dab(7)], shows that this hydrophobic domain is involved in LPS binding. The significant role of the positive charges is demonstrated with sp-Dap analogue, where L-alpha,gamma-diaminobutyric acid residues Dab(1) and Dab(8) are replaced by L-alpha,gamma-diaminopropionic acid (Dap), resulting in lower degrees of insertion in both LPS and PG monolayers. The importance of the N-terminal acyl chain is demonstrated with polymyxin B nonapeptide (PxB-np). PxB-np shows lower affinity for LPS compared to PxB, sp-B, or sp-C, but it does not insert into PG monolayers, although it binds superficially to the anionic film. Since PxB microbial killing appears to be mediated by osmotic instability due to OM-IM phospholipid exchange, the ability of the different peptides to induce membrane-membrane lipid exchange has been studied by use of phospholipid unilamellar vesicles. Results indicate that cationic amphipathicity determines peptide activity.