摘要
目的:研究抗菌肽Thanatin同系物(S-thanatin,Ts)的作用机制以及靶位。方法:进行Ts与脂多糖(LPS)的结合作用实验,用不同组分的磷脂制备出包裹了钙黄绿素的脂质体来模拟多肽与细胞膜的作用,另外研究多肽是否可以促使细胞内β-半乳糖苷酶的释放来指示细胞膜通透性的变化,用电子显微镜观察细菌在受抗菌肽作用后超微结构的改变。结果:Ts与LPS具有较强的亲和力(50%效应浓度为50μg·ml-1),多肽对脂质体有较强的扰乱作用,胆固醇的存在一定程度上抑制了这一作用。电镜图片显示未加药作用的菌体细胞膜边缘比较清晰圆滑,而被多肽作用的菌体细胞膜非常不规则,并且有破裂和内容物的释放。结论:初步阐明多肽Ts是通过作用于磷脂扰乱细胞膜发挥杀菌作用,并且很可能是通过静电作用与LPS结合。
Objective：To investigate the antibacterial mechanism of thanatin analog,antimicrobial peptide Sthanatin（Ts）.Methods：In this study,we investigated its lipopolysaccharide（LPS）-binding and neutralizing activity in vitro.We also investigated the effect of the presence of cholesterol on the interactions of the antimicrobial peptide Ts with phosphatidylcholine（PC） model membrane systems.Diffusion of extracellular o-nitrophenyl-β-Dgalactopyranoside（ONPG） into the cytoplasm of E.coli cells was monitored by measuring the production of o-nitrophenol.To further characterize the antimicrobial activity of Ts,transmission electron microscope（TEM） was employed to confirm morphological changes.Results：The results showed that Ts completely bound to the LPS（its median effective concentration being 50 μg·ml-1）.Membrane phospholipid was found to be the target for Ts in calcein leakage assay.Ts had changed the permeability of the membrane.Contrary to negative control,Ts produced chaotic membrane morphology and cell debris in electron microscopic appearance.Conclusion：These results supported our hypothesis that Ts first bound to negatively charged LPS and anionic lipid,and then disturbed the cytoplasmic membrane.This caused damage to the biological function and integrity of membrane.As a subsequence,the intracellular potential was exhausted and periplasmic material was released out of cells,which led to the doom of cells.
出处
《东南大学学报（医学版）》
CAS
2010年第4期423-427,共5页
Journal of Southeast University(Medical Science Edition)
基金
江苏省自然科学基金资助项目(BK2009274)
关键词
抗菌肽
S-thanatin
磷脂
细胞膜
作用机制
antimicrobial peptide
S-thanatin
phospholipid
cell membrane
mechanism of action
