Department of Biological Science, Ajou University, Suwon 442-749, Korea ; School of Biotechnology, Yeungnam University, Gyengsan 712-749, Korea.
School of Biotechnology, Yeungnam University, Gyengsan 712-749, Korea.
Plant Pathol J. 2014 Sep;30(3):245-53. doi: 10.5423/PPJ.OA.02.2014.0011.
Antimicrobial peptides (AMPs) are small but effective cationic peptides with variable length. In previous study, four hexapeptides were identified that showed antimicrobial activities against various phytopathogenic bacteria. KCM21, the most effective antimicrobial peptide, was selected for further analysis to understand its modes of action by monitoring inhibitory effects of various cations, time-dependent antimicrobial kinetics, and observing cell disruption by electron microscopy. The effects of KCM21 on Gram-negative strain, Pseudomonas syringae pv. tomato DC3000 and Gram-positive strain, Clavibacter michiganensis subsp. michiganensis were compared. Treatment with divalent cations such as Ca(2+) and Mg(2+) inhibited the bactericidal activities of KCM21 significantly against P. syringae pv. tomato DC3000. The bactericidal kinetic study showed that KCM21 killed both bacteria rapidly and the process was faster against C. michiganensis subsp. michiganensis. The electron microscopic analysis revealed that KCM21 induced the formation of micelles and blebs on the surface of P. syringae pv. tomato DC3000 cells, while it caused cell rupture against C. michiganensis subsp. michiganensis cells. The outer membrane alteration and higher sensitivity to Ca(2+) suggest that KCM21 interact with the outer membrane of P. syringae pv. tomato DC3000 cells during the process of killing, but not with C. michiganensis subsp. michiganensis cells that lack outer membrane. Considering that both strains had similar sensitivity to KCM21 in LB medium, outer membrane could not be the main target of KCM21, instead common compartments such as cytoplasmic membrane or internal macromolecules might be a possible target(s) of KCM21.
抗菌肽（AMPs）是具有可变长度的小而有效的阳离子肽。在以前的研究中，鉴定了四种六肽，它们对各种植物病原菌具有抗菌活性。选择最有效的抗菌肽 KCM21 进行进一步分析，通过监测各种阳离子的抑制作用、时间依赖性抗菌动力学以及观察电子显微镜下的细胞破裂来了解其作用模式。比较了 KCM21 对革兰氏阴性菌株丁香假单胞菌 pv.番茄 DC3000 和革兰氏阳性菌株密歇根棒状杆菌亚种密歇根的影响。二价阳离子如 Ca(2+)和 Mg(2+)的处理显著抑制了 KCM21 对 P. syringae pv.番茄 DC3000 的杀菌活性。杀菌动力学研究表明，KCM21 迅速杀死两种细菌，对 C. michiganensis subsp. michiganensis 的过程更快。电子显微镜分析显示，KCM21 在 P. syringae pv.番茄 DC3000 细胞表面诱导胶束和泡囊的形成，而对 C. michiganensis subsp. michiganensis 细胞则导致细胞破裂。外膜改变和对 Ca(2+)的更高敏感性表明，KCM21 在杀死过程中与 P. syringae pv.番茄 DC3000 细胞的外膜相互作用，但不与缺乏外膜的 C. michiganensis subsp. michiganensis 细胞相互作用。考虑到两种菌株在 LB 培养基中对 KCM21 的敏感性相似，外膜不可能是 KCM21 的主要靶标，相反，细胞质膜或内部大分子等常见隔室可能是 KCM21 的靶标（多个）。
