Localization and functional analysis of PepI, the immunity peptide of Pep5-producing Staphylococcus epidermidis strain 5
Localization and functional analysis of PepI, the immunity peptide of Pep5-producing Staphylococcus epidermidis strain 5
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DOI：
10.1128/aem.70.6.3263-3271.2004
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发表时间：
2004-06-01
影响因子：
4.4
通讯作者：
Sahl, HG
中科院分区：
文献类型：
--
作者：
Hoffmann, A;Schneider, T;Sahl, HG
Pep5 是一种由表皮葡萄球菌菌株 5 产生的阳离子成孔羊毛硫抗生素。生产菌株通过 69 个氨基酸的免疫肽 PepI 保护自身免受自身细菌素的致命作用。 PepI 的 N 端片段包含一段 20 个氨基酸的非极性残基，而 C 端非常亲水，带有净正电荷。我们使用绿色荧光蛋白 (GFP)-PepI 融合来获取其体内定位的信息。研究发现 PepI 存在于细胞质外，并在膜-细胞壁界面处积聚。细胞外定位似乎对于赋予免疫力至关重要。我们通过构建各种突变肽（也与 GFP 融合）来分析特定片段的功能作用。当通过引入带电氨基酸破坏 PepI 的疏水性 N 端片段时，PepI 的输出被阻断，并且表达此类突变肽的克隆对 Pep5 敏感。相反，当 PepI 在 C 端连续缩短时，其输出特性保持不变，但其赋予免疫力的能力逐渐降低。结果表明，N端部分是PepI转运所必需的，而C端部分对于赋予免疫表型很重要。针对PepI免疫机制提出了基于目标屏蔽的概念。
Pep5 is a cationic pore-forming lantibiotic produced by Staphylococcus epidermidis strain 5. The producer strain protects itself from the lethal action of its own bacteriocin through the 69-amino-acid immunity peptide PepI. The N-terminal segment of PepI contains a 20-amino-acid stretch of apolar residues, whereas the C terminus is very hydrophillic, with a net positive charge. We used green fluorescent protein (GFP)-PepI fusions to obtain information on its localization in vivo. PepI was found to occur outside the cytoplasm and to accumulate at the membrane-cell wall interface. The extracellular localization appeared essential for conferring immunity. We analyzed the functional role of the specific segments by constructing various mutant peptides, which were also fused to GFP. When the hydrophobic N-terminal segment of PepI was disrupted by introducing charged amino acids, the export of PepI was blocked and clones expressing such mutant peptides were Pep5 sensitive. When PepI was successively shortened at the C terminus, in contrast, its export properties remained unchanged whereas its ability to confer immunity was gradually reduced. The results show that the N-terminall part is required for the transport of PepI and that the C-terminal part is important for conferring the immunity phenotype. A concept based on target shielding is proposed for the PepI immunity mechanism.