Ferreira Ana Rita, Ferreira Mariana, Nunes Cláudia, Reis Salette, Teixeira Cátia, Gomes Paula, Gameiro Paula
LAQV/REQUIMTE (Laboratório Associado para a Química Verde-Rede de Química e Tecnologia), Departamento de Química e Bioquímica, Faculdade de Ciências, Universidade do Porto, Rua do Campo Alegre, s/n, 4169-007 Porto, Portugal.
LAQV/REQUIMTE, Laboratório de Química Aplicada, Faculdade de Farmácia da Universidade do Porto, Portugal, Rua de Jorge Viterbo Ferreira, 228, 4050-313 Porto, Portugal.
Membranes (Basel). 2023 Jan 21;13(2):138. doi: 10.3390/membranes13020138.
Cationic antimicrobial peptides (CAMPs) offer a promising strategy to counteract bacterial resistance, mostly due to their membrane-targeting activity. W-BP100 is a potent broad-spectrum cecropin-melittin CAMP bearing a single N-terminal Trp, which was previously found to improve its antibacterial activity. W-BP100 has high affinity toward anionic membranes, inducing membrane saturation at low peptide-to-lipid (P/L) ratios and membrane permeabilization, with the unique property of promoting the aggregation of anionic vesicles only at specific P/L ratios. Herein, we aimed to investigate this unusual behavior of W-BP100 by studying its aggregation and fusion properties with negatively-charged large (LUVs) or giant (GUVs) unilamellar vesicles using biophysical tools. Circular dichroism (CD) showed that W-BP100 adopted an α-helical conformation in anionic LUVs, neutralizing its surface charge at the aggregation P/L ratio. Its fusion activity, assessed by Förster resonance energy transfer (FRET) using steady-state fluorescence spectroscopy, occurred mainly at the membrane saturation/aggregation P/L ratio. Confocal microscopy studies confirmed that W-BP100 displays aggregation and detergent-like effects at a critical P/L ratio, above which it induces the formation of new lipid aggregates. Our data suggest that W-BP100 promotes the aggregation and fusion of anionic vesicles at specific P/L ratios, being able to reshape the morphology of GUVs into new lipid structures.
阳离子抗菌肽（CAMPs）为对抗细菌耐药性提供了一种很有前景的策略，这主要归功于它们的膜靶向活性。W-BP100是一种强效的广谱天蚕素-蜂毒肽CAMP，其N端带有一个色氨酸，此前发现该色氨酸可提高其抗菌活性。W-BP100对阴离子膜具有高亲和力，在低肽-脂（P/L）比时诱导膜饱和并导致膜通透性增加，具有仅在特定P/L比下促进阴离子囊泡聚集的独特性质。在此，我们旨在通过使用生物物理工具研究W-BP100与带负电荷的大单层囊泡（LUVs）或巨型单层囊泡（GUVs）的聚集和融合特性，来探究W-BP100的这种异常行为。圆二色性（CD）表明，W-BP100在阴离子LUVs中呈α螺旋构象，在聚集P/L比时中和其表面电荷。通过使用稳态荧光光谱的Förster共振能量转移（FRET）评估其融合活性，结果表明融合主要发生在膜饱和/聚集P/L比时。共聚焦显微镜研究证实，W-BP100在临界P/L比时表现出聚集和类似去污剂的作用，高于该比例时会诱导形成新的脂质聚集体。我们的数据表明，W-BP100在特定P/L比下促进阴离子囊泡的聚集和融合，能够将GUVs的形态重塑为新的脂质结构。
