UPMC Univ Paris 06, ER3 Biogenèse des Signaux Peptidiques (BIOSIPE), Paris, France.
PLoS One. 2013 Aug 13;8(8):e70782. doi: 10.1371/journal.pone.0070782. eCollection 2013.
Transcriptomic and peptidomic analysis of skin secretions from the Painted-belly leaf frog Phyllomedusa sauvagii led to the identification of 5 novel phylloseptins (PLS-S2 to -S6) and also of phylloseptin-1 (PSN-1, here renamed PLS-S1), the only member of this family previously isolated in this frog. Synthesis and characterization of these phylloseptins revealed differences in their antimicrobial activities. PLS-S1, -S2, and -S4 (79-95% amino acid sequence identity; net charge = +2) were highly potent and cidal against Gram-positive bacteria, including multidrug resistant S. aureus strains, and killed the promastigote stage of Leishmania infantum, L. braziliensis and L. major. By contrast, PLS-S3 (95% amino acid identity with PLS-S2; net charge = +1) and -S5 (net charge = +2) were found to be almost inactive against bacteria and protozoa. PLS-S6 was not studied as this peptide was closely related to PLS-S1. Differential scanning calorimetry on anionic and zwitterionic multilamellar vesicles combined with circular dichroism spectroscopy and membrane permeabilization assays on bacterial cells indicated that PLS-S1, -S2, and -S4 are structured in an amphipathic α-helix that disrupts the acyl chain packing of anionic lipid bilayers. As a result, regions of two coexisting phases could be formed, one phase rich in peptide and the other lipid-rich. After reaching a threshold peptide concentration, the disruption of lipid packing within the bilayer may lead to local cracks and disintegration of the microbial membrane. Differences in the net charge, α-helical folding propensity, and/or degree of amphipathicity between PLS-S1, -S2 and -S4, and between PLS-S3 and -S5 appear to be responsible for their marked differences in their antimicrobial activities. In addition to the detailed characterization of novel phylloseptins from P. sauvagii, our study provides additional data on the previously isolated PLS-S1 and on the mechanism of action of phylloseptins.
转录组和皮肤分泌物的肽组分析表明，从彩蛙 Phyllomedusa sauvagii 中鉴定出 5 种新型的 Phylloseptin（PLS-S2 到 S6），以及该蛙种中唯一分离出的 Phylloseptin-1（PSN-1，现更名为 PLS-S1）。这些 Phylloseptin 的合成和特性分析表明，它们的抗菌活性存在差异。PLS-S1、S2 和 S4（氨基酸序列同一性为 79-95%；净电荷 = +2）对革兰氏阳性菌具有很强的杀菌活性，包括耐多药的金黄色葡萄球菌菌株，并且可以杀死利什曼原虫的前鞭毛体阶段，L. braziliensis 和 L. major。相比之下，PLS-S3（与 PLS-S2 的氨基酸同一性为 95%；净电荷 = +1）和 PLS-S5（净电荷 = +2）对细菌和原生动物几乎没有活性。没有对 PLS-S6 进行研究，因为该肽与 PLS-S1 密切相关。阴离子和两性离子多层囊泡的差示扫描量热法结合圆二色性光谱和细菌细胞的膜通透性测定表明，PLS-S1、S2 和 S4 在两性α-螺旋中具有结构，破坏了阴离子脂质双层的酰基链堆积。结果，可以形成两个共存相的区域，一个富含肽的区域和另一个富含脂质的区域。在达到临界肽浓度后，双层内脂质堆积的破坏可能导致局部出现裂缝并破坏微生物膜。PLS-S1、S2 和 S4 之间以及 PLS-S3 和 S5 之间的净电荷、α-螺旋折叠倾向和/或两亲性的差异似乎是导致它们抗菌活性显著差异的原因。除了详细表征来自 P. sauvagii 的新型 Phylloseptin 外，我们的研究还提供了关于先前分离出的 PLS-S1 以及 Phylloseptin 作用机制的附加数据。
