pVEC hydrophobic N-terminus is critical for antibacterial activity
pVEC hydrophobic N-terminus is critical for antibacterial activity
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DOI：
10.1002/psc.3083
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发表时间：
2018-06-01
影响因子：
2.1
通讯作者：
Ozkirimli, Elif
中科院分区：
文献类型：
--
作者：
Alaybeyoglu, Begum;Akbulut, Berna Sariyar;Ozkirimli, Elif
细胞穿透肽（CPP）通常被定义为它们具有被内化到真核细胞中而不引起永久性膜损伤以及改善货物输送的共同能力。许多 CPP 还具有足够强的抗菌作用，可以选择性地裂解受感染的哺乳动物培养物中的微生物。 pVEC 是一种源自钙粘蛋白的 CPP，能够转位到哺乳动物细胞中，并且还具有抗菌作用。结构-活性关系和序列比对研究表明，pVEC 的疏水性 N 末端 (LLIIL) 对于该肽被真核细胞摄取至关重要。在本研究中，我们的目的是检查这些残留物对 pVEC 抗菌作用和易位机制的贡献。我们用 pVEC 和 del5 pVEC（pVEC 的 N 端截短变体）进行了抗菌活性和显微镜实验，结果表明，尽管这两种肽都会诱导膜通透性，但删除 LLIIL 残基后，pVEC 就会失去抗菌作用。我们还使用引导分子动力学模拟和复制交换伞采样模拟计算了运输过程的自由能，以比较两种肽在原子细节上的摄取机制的差异。尽管实验观察到的抗菌活性存在差异，但对 2 种肽的模拟显示出相似的特征，并且 pVEC 易位的能量消耗高于 del5 pVEC，表明 pVEC 摄取机制不能用简单的被动转运来解释。我们的结果表明，由于不可逆的膜破坏，LLIIL 残基是 pVEC 抗菌活性的关键贡献者。
Cell-penetrating peptides (CPPs) are commonly defined by their shared ability to be internalized into eukaryotic cells, without inducing permanent membrane damage, and to improve cargo delivery. Many CPPs also possess antimicrobial action strong enough to selectively lyse microbes in infected mammalian cultures. pVEC, a CPP derived from cadherin, is able to translocate into mammalian cells, and it is also antimicrobial. Structure-activity relationship and sequence alignment studies have suggested that the hydrophobic N-terminus (LLIIL) of pVEC is essential for this peptide's uptake into eukaryotic cells. In this study, our aim was to examine the contribution of these residues to the antimicrobial action and the translocation mechanism of pVEC. We performed antimicrobial activity and microscopy experiments with pVEC and with del5 pVEC (N-terminal truncated variant of pVEC) and showed that pVEC loses its antimicrobial effect upon deletion of the LLIIL residues, even though both peptides induce membrane permeability. We also calculated the free energy of the transport process using steered molecular dynamic simulations and replica exchange umbrella sampling simulations to compare the difference in uptake mechanism of the 2 peptides in atomistic detail. Despite the difference in experimentally observed antimicrobial activity, the simulations on the 2 peptides showed similar characteristics and the energetic cost of translocation of pVEC was higher than that of del5 pVEC, suggesting that pVEC uptake mechanism cannot be explained by simple passive transport. Our results suggest that LLIIL residues are key contributors to pVEC antibacterial activity because of irreversible membrane disruption.