In the present study, we characterized CF-14, a novel antimicrobial peptide derived from the catfish skin mucus. The objective of this study was to explore the antimicrobial mechanism of CF-14 against Escherichia coli. The agar-diffusion assay and the microdilution method were used to evaluate the antimicrobial activity and the minimum inhibitory concentration (MIC) of CF-14 against E. coli, respectively. In addition, the absorbance of the bacterial suspension filtrate at 260 nm was measured to quantify the leakage of bacterial cytoplasmic components. The bacterial morphological changes were observed by scanning electron microscopy, while confocal microscopy was used to investigate the localization site of CF-14 in E.coli. The DNA binding ability of CF-14 was evaluated using gel retardation assay and the binding of CF-14 to DnaK was evaluated using Discovery Studio. The results demonstrated that CF-14 exhibited strong antimicrobial activity against E.coli with an MIC of 31.3 μg/mL. Unlike common cationic anti-microbial peptides (AMPs) that target the cellmembrane, CF-14 penetrated the E.coli cell membrane and induced only minormembrane perturbations. Furthermore, the antimicrobial mechanism of CF-14 against E.coli involved DNA binding and competitive inhibition of bacterial DnaK. Finally, by deleting or replacing the amino acid sequence, the antibacterial activity of CF-14 was affected, which helped the optimization of amino acid sequence. Therefore, CF-14 can be a potential antimicrobial peptide.

中文翻译：

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细胞穿透肽CF-14对革兰氏阴性细菌大肠杆菌的抗菌活性和机理。

在本研究中，我们表征了CF-14，一种从from鱼皮肤粘液衍生的新型抗菌肽。这项研究的目的是探索CF-14对大肠杆菌的抗菌作用。琼脂扩散法和微稀释法分别用于评估CF-14对大肠杆菌的抗菌活性和最低抑菌浓度（MIC）。另外，测量细菌悬浮液滤液在260nm处的吸光度以定量细菌细胞质组分的泄漏。扫描电镜观察细菌的形态变化，共聚焦显微镜观察CF-14在大肠杆菌中的定位。CF-14的DNA结合能力使用凝胶阻滞法进行评估，CF-14与DnaK的结合使用Discovery Studio进行评估。结果表明，CF-14对大肠杆菌表现出强大的抗菌活性，MIC为31.3μg/ mL。与靶向细胞膜的常见阳离子抗微生物肽（AMPs）不同，CF-14穿透大肠杆菌细胞膜并仅引起较小的膜扰动。此外，CF-14对大肠杆菌的抗菌机制涉及DNA结合和竞争性抑制细菌DnaK。最后，通过删除或替换氨基酸序列，影响了CF-14的抗菌活性，从而有助于氨基酸序列的优化。因此，CF-14可能是潜在的抗菌肽。结果表明，CF-14对大肠杆菌表现出很强的抗菌活性，MIC为31.3μg/ mL。与靶向细胞膜的常见阳离子抗微生物肽（AMPs）不同，CF-14穿透大肠杆菌细胞膜并仅引起较小的膜扰动。此外，CF-14对大肠杆菌的抗菌机制涉及DNA结合和竞争性抑制细菌DnaK。最后，通过删除或替换氨基酸序列，影响了CF-14的抗菌活性，从而有助于氨基酸序列的优化。因此，CF-14可能是潜在的抗菌肽。结果表明，CF-14对大肠杆菌表现出强大的抗菌活性，MIC为31.3μg/ mL。与靶向细胞膜的常见阳离子抗微生物肽（AMPs）不同，CF-14穿透大肠杆菌细胞膜并仅引起较小的膜扰动。此外，CF-14对大肠杆菌的抗菌机制涉及DNA结合和竞争性抑制细菌DnaK。最后，通过删除或替换氨基酸序列，影响了CF-14的抗菌活性，从而有助于氨基酸序列的优化。因此，CF-14可能是潜在的抗菌肽。大肠杆菌细胞膜并仅引起微膜扰动。此外，CF-14对大肠杆菌的抗菌机制涉及DNA结合和竞争性抑制细菌DnaK。最后，通过删除或替换氨基酸序列，影响了CF-14的抗菌活性，从而有助于氨基酸序列的优化。因此，CF-14可能是潜在的抗菌肽。大肠杆菌细胞膜并仅引起微膜扰动。此外，CF-14对大肠杆菌的抗菌机制涉及DNA结合和竞争性抑制细菌DnaK。最后，通过删除或替换氨基酸序列，影响了CF-14的抗菌活性，从而有助于氨基酸序列的优化。因此，CF-14可能是潜在的抗菌肽。