Surface proteins and the formation of biofilms by Staphylococcus aureus
Surface proteins and the formation of biofilms by Staphylococcus aureus
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DOI：
10.1016/j.bbamem.2017.12.003
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发表时间：
2018-03-01
影响因子：
3.4
通讯作者：
Schaefer, Jacob
中科院分区：
文献类型：
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作者：
Kim, Sung Joon;Chang, James;Schaefer, Jacob
金黄色葡萄球菌生物膜作为持续性感染的储库造成严重的临床威胁。尽管这种临床意义，S。金黄色葡萄球菌生物膜是未知的。为了解决这些问题，我们使用固体核磁共振来检测S。金黄色葡萄球菌（SA 113），一株强生物膜形成菌。我们标记了静止期细胞的全细胞和细胞壁，在静止期后12-24 h形成年轻的生物膜，在静止期后60 h形成更成熟的生物膜。所有样品用（i）[N-15]甘氨酸和L-[1-C-13]苏氨酸标记，或在单独的实验中用（ii）L-[2-C-13，N-15]亮氨酸标记。然后，我们通过C{N}旋转回波双共振（REDOR）测量了C-13-N-15直接键。具有连接至表面蛋白的桥的肽聚糖茎的增加直接通过细胞壁双重差异（生物膜REDOR差异减去非质子REDOR差异）来确定。该方法消除了由N-15同位素富集的差异和C-13标记从苏氨酸降解到甘氨酸的路线所引起的误差。对于两种亲核细胞和成熟的生物膜，20%的五糖基桥是不交联的，是潜在的表面蛋白质附着位点。这些位点中没有一个具有附着在增殖细胞中的表面蛋白，但四分之一具有附着在成熟生物膜中的表面蛋白。此外，亮氨酸标记显示，在成熟的生物膜中，富含亮氨酸的区域中的β-链的浓度加倍。因此，建立S.金黄色葡萄球菌生物膜是用共价连接到细胞壁并促进细胞-细胞粘附的表面蛋白对细胞表面的广泛装饰。
Staphylococcus aureus biofilms pose a serious clinical threat as reservoirs for persistent infections. Despite this clinical significance, the composition and mechanism of formation of S. aureus biofilms are unknown. To address these problems, we used solid-state NMR to examine S. aureus (SA113), a strong biofilm-forming strain. We labeled whole cells and cell walls of planktonic cells, young bitifilms formed for 12-24 h after stationary phase, and more mature biofilms formed for up to 60 h after stationary phase. All samples were labeled either by (i) [N-15]glycine and L-[1-C-13] threonine, or in separate experiments, by (ii) L-[2-C-13,N-15]leucine. We then measured C-13-N-15 direct bonds by C{N} rotational-echo double resonance (REDOR). The increase in peptidoglycan stems that have bridges connected to a surface protein was determined directly by a cell-wall double difference (biofilm REDOR difference minus planktonic REDOR difference). This procedure eliminates errors arising from differences in N-15 isotopic enrichments and from the routing of C-13 label from threonine degradation to glycine. For both planktonic cells and the mature biofilm, 20% of pentaglycyl bridges are not cross-linked and are potential surface-protein attachment sites. None of these sites has a surface protein attached in the planktonic cells, but one-fourth have a surface protein attached in the mature biofilm. Moreover, the leucine-label shows that the concentration of beta-strands in leucine-rich regions doubles in the mature biofilm. Thus, a primary event in establishing a S. aureus biofilm is extensive decoration of the cell surface with surface proteins that are linked covalently to the cell wall and promote cell-cell adhesion.