Daptomycin is an acidic, 13-amino acid, cyclic polypeptide that contains a number of nonproteinogenic residues and is modified at its N-terminus with a decanoyl chain. It has been in clinical use since 2003 against selected drug-resistant Staphylococcus aureus and Enterococcus spp infections. In vitro, daptomycin is active against Gram-positive pathogens at low concentrations but its antibiotic activity depends critically on the presence of calcium ions. This dependence has been thought to arise from binding of one or two Ca²⁺ ions to daptomycin as a required step in its interaction with the bacterial membrane. Here, we investigated the interaction of daptomycin with giant unilamellar vesicles (GUVs) composed 1-palmitoyl-2-oleoylphosphatidylcholine (POPC) and 1-palmitoyl-2-oleoylphosphatidylglycerol (POPG). We used fluorescence confocal microscopy to monitor binding of the peptide to GUVs and follow its effect on the membrane of the vesicle. We found that in the absence of POPG or Ca²⁺ daptomycin does not bind measurably to the lipid membrane. In the presence of 20–30% PG in the membrane and 2 mM Ca²⁺, daptomycin induces the formation of membrane domains rich in acidic lipids. This effect is not induced by Ca²⁺ alone. In addition, daptomycin causes GUV collapse, but it does not translocate across the membrane to the inside of intact POPC/POPG vesicles. We conclude that pore formation is probably not the mechanism by which the peptide functions. On the other hand, we found that daptomycin coclusters with the anionic phospholipid POPG and the fluorescent probes used, leading to extensive formation of daptomycin–POPG domains in the membrane.

中文翻译：

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脂质膜中的达托霉素-磷脂酰甘油域

达托霉素是一种酸性的13个氨基酸的环状多肽，含有许多非蛋白残基，并在其N端被癸酰基链修饰。自2003年以来，它已在临床上针对选定的耐药性金黄色葡萄球菌和肠球菌属感染。在体外，达托霉素在低浓度下对革兰氏阳性病原体具有活性，但其抗生素活性主要取决于钙离子的存在。人们认为这种依赖性是由一个或两个Ca ^2+的结合引起的。离子达托霉素是其与细菌膜相互作用的必需步骤。在这里，我们研究了达托霉素与组成1-棕榈酰基-2-油酰基磷脂酰胆碱（POPC）和1-棕榈酰基-2-油酰基磷脂酰甘油（POPG）的单层囊泡（GUV）的相互作用。我们使用荧光共聚焦显微镜来监测肽与GUV的结合，并跟踪其对囊泡膜的影响。我们发现，在不存在POPG或Ca ²⁺的情况下，达托霉素不能与脂质膜可测量地结合。在膜中存在20–30％PG和2 mM Ca ^{2+的情况下}，达托霉素可诱导形成富含酸性脂质的膜结构域。Ca ²⁺不会诱导这种作用独自的。此外，达托霉素会引起GUV崩溃，但不会跨膜移位到完整的POPC / POPG囊泡内部。我们得出的结论是，孔的形成可能不是肽发挥作用的机制。另一方面，我们发现达托霉素与阴离子磷脂POPG和所用的荧光探针紧密结合，导致膜中达托霉素-POPG结构域大量形成。