源自蜡样芽孢杆菌抗菌肽DB16的筛选及其对金黄色葡萄球菌的抑菌机制  

Selection of Antimicrobial Peptide DB16 from Bacillus cereus and Its Antibacterial Mechanism against Staphylococcus aureus

作　　者：杨智源 金日天 梁铎 邱绪建[1,2] 杨燊 林蓉[1,2] YANG Zhiyuan;JIN Ritian;LIANG Duo;QIU Xujian;YANG Shen;LIN Rong(College of Ocean Food and Biological Engineering,Jimei University,Xiamen 361021,China;Collaborative Innovation Center for Key Technologies of Deep Processing of Marine Food,Dalian Polytechnic University,Dalian 116034,China;Fujian Key Laboratory of Food Microbiology and Enzyme Engineering,Xiamen 361021,China)

机构地区：[1]集美大学海洋食品与生物工程学院,福建厦门361021 [2]海洋食品精深加工关键技术省部共建协同创新中心,大连工业大学,辽宁大连116034 [3]福建省食品微生物与酶工程重点实验室,福建厦门361021

出　　处：《食品科学》2024年第10期126-134,共9页Food Science

基　　金：福建省高校产学合作项目(2023N5008)。

摘　　要：本实验发现当与蜡样芽孢杆菌混合培养时,金黄色葡萄球菌的生长受到抑制。结合生物信息学方法从蜡样芽孢杆菌DeadBoxATP依赖性RNA解旋酶中预测、筛选得到抗菌肽DB16(RKLLQFAKKLGIVFTK)。抗菌肽DB16对金黄色葡萄球菌的最低抑菌质量浓度(minimum inhibitory concentration,MIC)为31.25μg/mL,可在0.5 h内完全抑制金黄色葡萄球菌的生长;抗菌肽DB16在磷酸盐缓冲溶液中呈现无规卷曲结构,在十二烷基硫酸钠环境中转变为α-螺旋结构。采用荧光探针、流式细胞术、透射电子显微镜、DNA凝胶电泳以及圆二色谱等方法探究抗菌肽DB16对金黄色葡萄球菌的抑菌机制,结果表明,抗菌肽DB16能够改变细菌细胞膜通透性,造成细胞内容物外泄,同时进入胞内与金黄色葡萄球菌基因组DNA结合,影响正常DNA的复制,最终抑制菌体的生长繁殖。此外,对哺乳动物红细胞的处理表明,DB16在8×MIC条件下仍无溶血性。综上,源自蜡样芽孢杆菌的抗菌肽DB16在金黄色葡萄球菌防控方面具有很大应用潜力。In this study,it was found that when co-cultured with Bacillus cereus,the growth of Staphylococcus aureus was inhibited.The antimicrobial peptide DB16(RKLLQFAKKLGIVFTK)was predicted and selected from the ATP-dependent RNA helicase of B.cereus by bioinformatics.The minimum inhibitory concentration(MIC)of the antimicrobial peptide DB16 against S.aureus was 31.25μg/mL,which could completely inhibit the growth of S.aureus within 0.5 h.DB16 presented a random coiled structure in phosphate buffered saline(PBS),and turned into anα-helix structure in sodium dodecyl sulfate(SDS).Fluorescence probe,flow cytometry,transmission electron microscopy(TEM),DNA gel electrophoresis and circular dichroism(CD)spectroscopy were used to explore the antibacterial mechanism of DB16 against S.aureus.The results showed that the antibacterial peptide DB16 could change the permeability of the bacterial cell membrane,thereby causing the leakage of cellular contents,while entering the cells to combine with the genomic DNA of S.aureus,affecting normal DNA replication,and finally inhibiting the growth and reproduction of bacteria.In addition,DB16 at 8×MIC showed no hemolysis on mammalian red blood cells.In conclusion,the antimicrobial peptide DB16 from B.cereus has great potential for the prevention and control of S.aureus.

关 键 词：蜡样芽孢杆菌 金黄色葡萄球菌 抗菌肽 细胞膜通透性 DNA结合机制 

分 类 号：Q939.92[生物学—微生物学]