INTERACTION OF HUMAN DEFENSINS WITH ESCHERICHIA-COLI - MECHANISM OF BACTERICIDAL ACTIVITY
INTERACTION OF HUMAN DEFENSINS WITH ESCHERICHIA-COLI - MECHANISM OF BACTERICIDAL ACTIVITY
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DOI：
10.1172/jci114198
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发表时间：
1989-08-01
影响因子：
15.9
通讯作者：
SELSTED, ME
中科院分区：
文献类型：
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作者：
LEHRER, RI;BARTON, A;SELSTED, ME
防御素是一种小的、富含半胱氨酸的抗菌肽，在人、兔和豚鼠的中性粒细胞(PMN)中含量丰富。三种防御素(人中性粒细胞多肽防御素[HNP]-1、HNP-2和HNP-3)占人PMN天青颗粒总蛋白的30%~50%。我们研究了HNP介导的对大肠杆菌ML-35(i-，y-，z+)及其pBR322转化的衍生物E.coliML-35P的杀菌作用机制。在支持杀菌活性的条件下，HNP-1依次对大肠杆菌的外膜(OM)和内膜(IM)进行渗透。与这些事件同时发生的是，细菌对DNA、RNA和蛋白质的合成停止，菌落数量下降。尽管这些事件在标准化验条件下紧密耦合。当用甘露醇质溶化的大肠杆菌进行实验时，OM通透性与IM通透性部分分离。在这样的条件下，细菌死亡的速度和程度与IM完整性的丧失比OM通透性的丧失更密切地平行。被防御素杀死的大肠杆菌的电子显微镜显示，在周质空间中存在着惊人的电子致密沉积物，并附着在OM上。总体而言，这些研究表明，HNP介导的对大肠杆菌ML-35的杀菌活性与OM和IM的顺序通透性有关，内膜通透性似乎是致死事件。
Defensins are small, cysteine-rich antimicrobial peptides that are abundant in human, rabbit, and guinea pig neutrophils (PMN). Three defensins (human neutrophil peptide defensin [HNP]-1, HNP-2, and HNP-3) constitute between 30 and 50% of the total protein in azurophil granules of human PMN. We examined the mechanism of HNP-mediated bactericidal activity against Escherichia coli ML-35 (i-, y-, z+) and its pBR322-transformed derivative, E. coli ML-35p. Under conditions that supported bactericidal activity, HNP-1 sequentially permeabilized the outer membrane (OM) and inner membrane (IM) of E. coli. Coincident with these events, bacterial synthesis of DNA, RNA, and protein ceased and the colony count fell. Although these events were closely coupled under standard assay conditions. OM permeabilization was partially dissociated from IM permeabilization when experiments were performed with E. coli that had been plasmolyzed by mannitol. Under such conditions, the rate and extent of bacterial death more closely paralleled loss of IM integrity than OM permeabilization. Electron microscopy of E. coli that had been killed by defensins revealed the presence of striking electron-dense deposits in the periplasmic space and affixed to the OM. Overall, these studies show that HNP-mediated bactericidal activity against E. coli ML-35 is associated with sequential permeabilization of the OM and IM, and that inner membrane permeabilization appears to be the lethal event.