新型抗铜绿假单胞菌双功能杂合抗菌肽LLM的设计及特性研究  

Design and characterization of a bi-functional bybrid antibacterial peptide LLM against Pseudomonas aeruginosa

作　　者：张泽瑾 姚宏纪 薛兆毅 展羽桐 郭莹洁 吴蒙 蔺可鑫 熊婧妍 张勇 ZHANG Ze-jin;YAO Hong-ji;XUE Zhao-yi;ZHAN Yu-tong;GUO Ying-jie;WU Meng;LIN Ke-xin;XIONG Jing-yan;ZHANG Yong(The First Clinical College of Hainan Medical University,Haikou 571199,China;School of Basic Medicine and Life Science,Hainan Medical University,Haikou 571199,China)

机构地区：[1]海南医学院第一临床学院,海南海口571199 [2]海南医学院基础医学与生命科学学院,海南海口571199

出　　处：《海南医学院学报》2022年第21期1627-1633,共7页Journal of Hainan Medical University

基　　金：海南省高等学校科学研究资助项目(Hnky2019ZD-32);大学生创新创业训练计划项目(S202011810018)。

摘　　要：目的:设计抗铜绿假单胞菌的新型双功能(杀菌和中和内毒素)的杂合抗菌肽,并探究其杀菌特性。方法:拟以四肽GGGS-Linker偶联中和内毒素多肽和抗铜绿假单胞菌活性抗菌肽,构建新型双功能杂合抗菌肽,并对结构特征进行分析;测定其抗铜绿假单胞菌的活性和杀菌特点;进一步评价目标杂合肽对内毒素的中和作用和对兔红细胞的溶血性。结果:设计并获得具有高抗铜绿假单胞菌和中和内毒素双重功效的杂合抗菌肽LLM,其携带+8正电荷;LLM对铜绿假单胞菌P.aeruginosa CMCC10104和P.aeruginosa ATCC 9027的MIC分别为2、4μmol/L,MBC/MIC=1,其抗菌活性均优于母体肽;LLM能够快速杀菌,1 h处理能够杀灭99%以上的铜绿假单胞菌。LLM对内毒素呈现较强的中和作用,能够显著降低培养液中内毒素产生水平;LLM在高达115μmol/L(400μg/mL)时对兔红细胞没有表现出显著的溶血性。结论:LLM抗铜绿假单胞菌的MIC为2~4μmol/L,能够中和内毒素,可以快速杀菌,且浓度低于115μmol/L时未表现出显著溶血性。Objective:To design a bifunctional antimicrobial peptide with high antibacterial activity and endotoxin neutralization against P.aeruginosa and explore its bactericidal properties.Methods:The neutralizing endotoxin peptides and antimicrobial peptide against P.aeruginosa were connected by a GGGS-linker to get a bi-functional bybrid peptide.Its structural parameter was tested by EMBOSS software.The minimum inhibitory concentration(MIC),minimum bactericidal concentration(MBC)and bactericidal kinetics against P.aeruginosa were determined.Its effect on endotoxin neutralization and hemolysis were also evaluated.Results:We designed and obtained a bybrid peptide LLM,which carried+8 positive charge with high activity against P.aeruginosa and neutralizing endotoxin.The MIC of LLM against P.aeruginosa CMCC10104 and P.aeruginosa ATCC 9027 were 2and 4μmol/L,and MBC/MIC equal 1.LLM showed rapid anti-P.aeruginosa effects and significantly neutralized the endotoxin released,and not exhibited hemolysis as high as 115μmol/L(400μg/mL).Conclusion:The MICs of LLM against P.aeruginosa were 2~4μmol/L,it showed significant activity of anti-P.aeruginosa and neutralizing endotoxin,and could kill bacteria quickly,and did not show significant hemolytic under 115μmol/L.

关 键 词：杂合抗菌肽 铜绿假单胞菌 内毒素中和 杀菌 溶血性 

分 类 号：R914[医药卫生—药物化学]