Mechanism study on a new antimicrobial peptide Sphistin derived from the N-terminus of crab histone H2A identified in haemolymphs of Scylla paramamosain
Mechanism study on a new antimicrobial peptide Sphistin derived from the N-terminus of crab histone H2A identified in haemolymphs of Scylla paramamosain
复制标题
拟青蟹血淋巴中蟹组蛋白H2A N端新型抗菌肽Sphistin的作用机制研究
DOI：
10.1016/j.fsi.2015.10.010
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发表时间：
2015-12-01
影响因子：
4.7
通讯作者：
Wang, Ke-Jian
中科院分区：
文献类型：
--
作者：
Chen, Bei;Fan, Dan-Qing;Wang, Ke-Jian
组蛋白H2 A通过产生特异性抗菌肽（AMPS）参与宿主的免疫防御，因此对该类抗菌肽在脊椎动物和无脊椎动物中的特性进行研究已成为一个热点。尽管在各种生命物种中已经报道了数千种AMP，但在螃蟹中仅知道几种AMP，特别是尚未报道H2 A衍生的AMP。本研究通过对拟穴青蟹（Scylla paramamosain）组蛋白H2 A的序列分析，确定了一个由38个氨基酸组成的具有抗菌活性的多肽。组蛋白H2 A衍生肽是一种AMP样分子，命名为Sphistin。Sphistin具有典型的AMP特征，如两亲性α螺旋二级结构和正电荷网。合成的Sphistin对革兰氏阳性菌、革兰氏阴性菌和酵母菌均有较好的抗菌活性，其中嗜水气单胞菌、荧光假单胞菌和施氏假单胞菌是重要的水产病原菌。使用扫描电子显微镜在用Sphistin处理的细菌细胞中观察到细胞内容物的泄漏和细胞表面的破坏。证明了鞘氨醇引起的大肠杆菌细胞质膜透性增加。共聚焦显微镜下观察发现，Sphistin可以联合收割机到金黄色葡萄球菌、大肠杆菌等的膜上。coliMC 1061和毕赤酵母中表达，但不转移到细胞质中。此外，Sphistin与LPS或LTA的亲和力也证明了Sphistin与细胞膜之间存在相互作用。因此，该肽的抗微生物机制可能通过细菌细胞膜的吸附和随后的透化而不是穿透细胞膜来发挥作用。此外，即使在100 μ g/mL的高浓度下24小时，合成的鞘氨醇对原代培养的蟹血淋巴和哺乳动物细胞也没有显示出细胞毒性。这是首次从S.拟穴蛙肽具有特异的抗菌活性和作用机制，有望在水产养殖和兽医学中得到应用。(C)2015爱思唯尔有限公司版权所有。
Histone H2A is known to participate in host immune defense through generating special antimicrobial peptides (AMPS), for which it has been an interesting research focus to characterize this kind of peptides in vertebrates and invertebrates. Although thousands of AMPs have been reported in variety of life species, only several AMPs are known in crabs and in particular no H2A-derived AMP has yet been reported. In the present study, a 38-amino acid peptide with antimicrobial activity was determined based on the sequence analysis of a histone H2A identified from the mud crab Scylla paramamosain. The histone H2A derived peptide was an AMP-like molecule and designated as Sphistin. Sphistin showed typical features of AMPs such as amphiphilic alpha-helical second structrue and positive charge net. The synthetic Sphistin exerted high antimicrobial activity against Gram-positive, Gram-negative bacteria and yeast, among which Aeromonas hydrophila, Pseudomonas fluorescens and Pseudomonas stutzeri are important aquatic pathogens. Leakage of the cell content and disruption of the cell surface were observed in bacterial cells treated with Sphistin using scanning electron microscopy. It was proved that the increasing cytoplasmic membrane permeability of Escherichia coli was caused by Sphistin. Further observation under confocal microscopy showed that Sphistin could combine onto the membrane of Staphylococcus aureus, E. coli MC1061 and Pichia pastoris but not translocate into the cytoplasm. Moreover, the affinity of Sphistin with either LPS or LTA was also testified that there was an interaction between Sphistin and cell membrane. Thus, the antimicrobial mechanism of this peptide likely exerted via adsorption and subsequently permeabilization of the bacterial cell membranes other than penetrating cell membrane. In addition, synthetic Sphistin exhibited no cytotoxicity to primary cultured crab haemolymphs and mammalian cells even at a high concentration of 100 mu g/mL for 24 h. This is the first report of a histone-derived Sphistin identified from S. paramamosain with a specific antimicrobial activity and mechanism, which could be a new candidate for future application in aquaculture and veterinary medicine. (C) 2015 Elsevier Ltd. All rights reserved.