乳源抗菌肽BCp12对金黄色葡萄球菌蛋白修饰介导的抗菌作用分子机制

致病菌蛋白修饰是抑菌机制的研究热点，但主要集中在酸、碱、盐、氧、金属离子等使致病菌发生蛋白修饰影响其生长代谢的研究，而蛋白修饰介导的抗菌肽抑菌机理的研究鲜见报道。利用前期项目筛选的乳源抗菌肽BCp12和金葡菌DC.RB_015为材料，通过分析抗菌肽与菌体壁膜亲和作用、泛抗体结合Western Blot分析、全蛋白TMT标记定量蛋白组学、LC-MS/MS及差异修饰蛋白定量分析，研究抗菌肽对金葡菌蛋白修饰水平的影响、差异修饰蛋白及位点；依据金葡菌生长代谢途径，基于蛋白修饰互作网络、修饰位点保守性和功能域分析，研究被修饰蛋白参与的代谢通路以及差异表达的修饰酶，筛选3-4个关键修饰靶点；通过靶标蛋白PRM定量分析和点突变实验，验证蛋白修饰的关键靶点，揭示蛋白修饰介导的抑菌机制。项目为功能型乳源抗菌肽、天然食品防腐剂及抗生素替代品的研制提供科学依据，为云南特色乳产业发展和21世纪大健康产业服务。

In recent years, protein modification of pathogenic bacteria has been a central research issue in the bacteriostasis mechanism. However, it mainly focuses on the protein modification factors of such as acid, alkali, salt, oxygen, and metal ions, to affect their growth and reproduction. There are only a few reports on the mechanism of antibacterial peptides mediated by protein modification. In this current work, the milk-derived antimicrobial peptide BCp12 and S. aureus DC.RB_015 screened from the previous project selected as the primary material. The differential protein and whole protein TMT labeling quantitative proteomics elucidated by outlining the types of protein modifications that affect the growth and reproduction of S. aureus. Those parameters analyzed by determining the affinity between the antimicrobial peptide and bacterial wall membrane, pan-antibodies binding Western Blot, LC-MS/MS, and quantitative analysis of the differentially modified proteins. Modification of proteins and sites created based on the growth and metabolic pathway of S. aureus, protein-protein interaction network, conservatism site, and functional domain analysis, and also the study of the metabolic pathways and differentially expressed modified metabolic enzymes involved in the modified protein. The critical modification targets of this work pointed to obtain the experimental results of quantitative analysis of the protein PRM target and mutation sites, verify the key points of protein modification, and reveal the mechanism of protein modification affecting bacterial metabolism. The work provides a scientific basis for the development of functional milk-derived antibacterial peptides, natural food preservatives, and antibiotic substitutes. It also supports the improvement of Yunnan’s typical dairy industry and the 21st-century health industry as well.