Department of Systems Biology, Technical University of Denmark, Kgs. Lyngby, Denmark.
PLoS One. 2013 Jul 12;8(7):e69483. doi: 10.1371/journal.pone.0069483. Print 2013.
The peptidomimetic LTX109 (arginine-tertbutyl tryptophan-arginine-phenylethan) was previously shown to have antibacterial properties. Here, we investigated the activity of this novel antimicrobial peptidomimetic on the yeast Saccharomyces cerevisiae. We found that LTX109 was an efficient fungicide that killed all viable cells in an exponentially growing population as well as a large proportion of cells in biofilm formed on an abiotic surface. LTX109 had similar killing kinetics to the membrane-permeabilizing fungicide amphotericin B, which led us to investigate the ability of LTX109 to disrupt plasma membrane integrity. S. cerevisiae cells exposed to a high concentration of LTX109 showed rapid release of potassium and amino acids, suggesting that LTX109 acted by destabilizing the plasma membrane. This was supported by the finding that cells were permeable to the fluorescent nucleic acid stain SYTOX Green after a few minutes of LTX109 treatment. We screened a haploid S. cerevisiae gene deletion library for mutants resistant to LTX109 to uncover potential molecular targets. Eight genes conferred LTX109 resistance when deleted and six were involved in the sphingolipid biosynthetic pathway (SUR1, SUR2, SKN1, IPT1, FEN1 and ORM2). The involvement of all of these genes in the biosynthetic pathway for the fungal-specific lipids mannosylinositol phosphorylceramide (MIPC) and mannosyl di-(inositol phosphoryl) ceramide (M(IP)2C) suggested that these lipids were essential for LTX109 sensitivity. Our observations are consistent with a model in which LTX109 kills S. cerevisiae by nonspecific destabilization of the plasma membrane through direct or indirect interaction with the sphingolipids.
此前有研究表明，拟肽 LTX109（精氨酸叔丁基色氨酸精氨酸苯乙）具有抗菌特性。在这里，我们研究了这种新型抗菌拟肽对酵母酿酒酵母的活性。我们发现 LTX109 是一种有效的杀菌剂，它可以杀死指数生长期的所有活细胞，以及在非生物表面形成的生物膜中的很大一部分细胞。LTX109 的杀菌动力学与膜通透杀菌剂两性霉素 B 相似，这促使我们研究 LTX109 破坏质膜完整性的能力。暴露于高浓度 LTX109 的酿酒酵母细胞迅速释放钾和氨基酸，表明 LTX109 通过破坏质膜的稳定性起作用。几分钟后，细胞对荧光核酸染料 SYTOX Green 具有通透性，这一发现支持了这一观点。我们筛选了一个单倍体酿酒酵母基因缺失文库，以寻找对 LTX109 具有抗性的突变体，以揭示潜在的分子靶标。当缺失八个基因时，赋予了 LTX109 抗性，其中六个基因参与了鞘脂生物合成途径（SUR1、SUR2、SKN1、IPT1、FEN1 和 ORM2）。所有这些基因都参与了真菌特异性脂质甘露糖肌醇磷酸神经酰胺（MIPC）和甘露糖二（肌醇磷酸）神经酰胺（M（IP）2C）的生物合成途径，表明这些脂质对 LTX109 的敏感性至关重要。我们的观察结果与一种模型一致，即 LTX109 通过与鞘脂的直接或间接相互作用，通过非特异性破坏质膜的稳定性来杀死酿酒酵母。
