该研究旨在评估驼乳源乳铁蛋白(lactoferrampin-lactoferricin, LFA-LFC)嵌合肽对变异链球菌、唾液链球菌和远缘链球菌的抗菌效果并初步探究其作用机制。首先预测并鉴定驼乳源LFA-LFC嵌合肽的分子特性和二级结构;通过测定嵌合肽最低抑菌(minimum inhibitory concentration, MIC)、杀菌浓度(minimum bactericidal concentration, MBC),绘制时间-致死曲线,评估嵌合肽的抗菌效果;分析嵌合肽对生物膜的预防、消除作用。采用电子显微镜观察嵌合肽对细菌的作用位点,测定菌体DNA迁移率,对嵌合肽的抗菌机制进行初步研究。结果表明,驼乳源LFA-LFC嵌合肽对变异链球菌、唾液链球菌和远缘链球菌的最低抑菌浓度分别为32、32、64 μmol/L;最低杀菌浓度分别为128、128、256 μmol/L;细菌在1×MIC嵌合肽浓度处理30 min后完全致死;当嵌合肽浓度为128 μmol/L时预防生物膜生成约80%,512 μmol/L时消除生物膜约50%;4×MIC嵌合肽浓度处理细菌后发现其细胞膜均出现裂解,DNA未发生迁移。该研究认为驼乳源LFA-LFC嵌合肽对3株菌表现出显著的抗菌效果,嵌合肽可通过破坏细菌细胞膜,进入细胞内部作用于DNA,抑制细菌生长繁殖。
The aim of this study was to evaluate the antibacterial effect of camel milk-derived lactoferrampin-lactoferricin (LFA-LFC) chimeric peptides against Streptococcus mutans, Streptococcus salivarius, and Streptococcus sobrinus and to investigate the mechanism of action. The molecular properties and secondary structure of the camel milk LFA-LFC chimeric peptide were predicted and characterized. The antibacterial effect of the chimeric peptide was evaluated by determining the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) and plotting the time-lethal curve. The preventive and eliminative effects of the chimeric peptide on biofilm was determined. The action sites of chimeric peptide on bacteria were observed by electron microscopy, and the DNA migration rate of the bacteria was measured. The results showed that MIC of LFA-LFC chimeric peptide were 32, 32, and 64 μmol/L for S. mutans, S. salivarius and S. salivarius, respectively. The MBC were 128, 128, and 256 μmol/L, respectively. The bacteria were lethal after 30 min treated with 1×MIC chimeric peptide concentration. When the chimeric peptide concentration was 128 μmol/L, 80% of biofilm formation was prohibited, and at 512 μmol/L, about 50% of biofilm were eliminated. When 4×MIC chimeric peptide concentrations were used to treat the bacteria, all cell membranes were cleaved and no DNA migration occurred. The camel milk-derived LFA-LFC chimeric peptide showed significant antibacterial effects against the three strains of bacteria; the chimeric peptide could inhibit the growth and reproduction of bacteria by disrupting the bacterial cell membrane and entering the cell to act on the DNA.
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