Proteomic profiling of cell envelope-associated proteins from Staphylococcus aureus
Proteomic profiling of cell envelope-associated proteins from Staphylococcus aureus
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DOI：
10.1002/pmic.200500253
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发表时间：
2006-03-01
期刊：
影响因子：
3.4
通讯作者：
Peterson, SN
中科院分区：
文献类型：
--
作者：
Gatlin, CL;Pieper, R;Peterson, SN
社区获得性金黄色葡萄球菌高毒力的出现以及对多种抗菌药物（包括甲氧西林和万古霉素）耐药的持续进展标志着S.金黄色葡萄球菌是一个严重的健康威胁。研究定位于细胞表面的蛋白质有助于阐明S.金黄色。在这项研究中，开发了蛋白质组学分析方法，以溶解、显示和评估溶葡萄球菌酶消化的来自培养的万古霉素中间体S的细胞包膜的上清液中存在的蛋白质的丰度水平。金黄色葡萄球菌（VISA）细胞。结合方法的2-DE或色谱分离的蛋白质与MS分析，导致在144个特别感兴趣的蛋白质的鉴定。在这些蛋白质中，48个含有预测的细胞壁定位或输出信号基序，包括14个具有不同共价肽聚糖锚定位点的蛋白质，其中4个至今尚未鉴定。两种最丰富的细胞包膜蛋白之一是S.金黄色葡萄球菌表面蛋白G许多分泌的蛋白质，不是共价细胞壁锚定的显示，表明，在指数生长期，分泌的蛋白质可以生理保留在细胞包膜，并可能与细胞壁锚定的蛋白质和碳水化合物结构的方式尚未确定。剩余的96种蛋白质，缺乏可识别的基序，在VISA细胞包膜组分中重复分析。我们描述了一种新的半定量方法，以确定丰富的因素，这样的蛋白质在2-DE凝胶的细胞包膜组分相对于全细胞裂解物，并讨论这些数据的背景下，真正的细胞包膜本地化与实验引起的细胞裂解。
The emergence of highly virulent community acquired Staphylococcus aureus and continued progression of resistance to multiple antimicrobials, including methicillin and vancomycin, marks the reemergence of S. aureus as a serious health care threat. Investigation of proteins localized to the cell surface could help to elucidate mechanisms of virulence and antibiotic resistance in S. aureus. In this study, proteomic profiling methods were developed to solubilize, display, and evaluate abundance levels of proteins present in the supernatants of the lysostaphin-digested cell envelope from cultured vancomycin-intermediate S. aureus (VISA) cells. Combining approaches of 2-DE or chromatographic separation of proteins with MS analyses resulted in the identification of 144 proteins of particular interest. Of these proteins, 48 contained predicted cell wall localization or export signal motifs, including 14 with distinct covalent peptidoglycan-anchor sites, four of which are uncharacterized to date. One of the two most abundant cell envelope proteins, which showed remarkably high variations in MW and pI in the 2-DE gel display, was the S. aureus surface protein G. The display of numerous secreted proteins that are not covalently cell wall-anchored, suggests that, in the exponential growth phase, secreted proteins can be retained physiologically in the cell envelope and may interact with cell wall-anchored proteins and carbohydrate structures in a manner yet to be determined. The remaining 96 proteins, devoid of recognizable motifs, were repeatedly profiled in the VISA cell envelope fractions. We describe a novel semiquantitative method to determine abundance factors of such proteins in 2-DE gels of cell envelope fractions relative to whole cell lysates and discuss these data in the context of true cell envelope localization versus experimentally caused cell lysis.