Simultaneous qualitative and quantitative analysis of the Escherichia coli Proteome -: A sweet tale
Simultaneous qualitative and quantitative analysis of the Escherichia coli Proteome -: A sweet tale
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DOI：
10.1074/mcp.m500321-mcp200
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发表时间：
2006-04-01
影响因子：
7
通讯作者：
Geromanos, SJ
中科院分区：
文献类型：
--
作者：
Silva, JC;Denny, R;Geromanos, SJ
我们描述了一种新的LC MS方法来相对定量和同时鉴定复杂环境中的未分离的大肠杆菌中的蛋白质。这种无标记的LC MS采集方法可以观察所有可检测的洗脱肽及其相应的碎片离子。采集后数据分析方法提取胰蛋白酶多肽的色谱学和质谱学信息，以提供时间分辨的、准确的质量测量，这些测量随后用于对组成蛋白质的定量和鉴定。大肠杆菌对碳源变化的反应是众所周知的，因此在验证分析方法时，它通常被用作模型生物系统。使用这种LC MS方法，我们鉴定了从生长在葡萄糖、乳糖和醋酸盐中的大肠杆菌中分离出的蛋白质。相应蛋白质的相对丰度的变化是从两种情况下共有的多肽测量的。对于那些对每个条件都是唯一的多肽，还确定了蛋白质的同一性，并且发现这些同一性与生长条件施加的潜在的生化限制是一致的。在三个二元比较中，特征蛋白质丰度的相对变化从0.1到90倍不等。特征蛋白的总覆盖率从10%到80%不等，每个蛋白由1到34个多肽组成。我们的研究获得的定量结果与其他现有的蛋白质组学和转录图谱方法相当。这项研究展示了这种新的LC MS方法在复杂混合物中蛋白质的同时定量和全面定性分析中的稳健性。
We describe a novel LCMS approach to the relative quantitation and simultaneous identification of proteins within the complex milieu of unfractionated Escherichia coli. This label-free, LCMS acquisition method observes all detectable, eluting peptides and their corresponding fragment ions. Postacquisition data analysis methods extract both the chromatographic and the mass spectrometric information on the tryptic peptides to provide time-resolved, accurate mass measurements, which are subsequently used for quantitation and identification of constituent proteins. The response of E. coli to carbon source variation is well understood, and it is thus commonly used as a model biological system when validating an analytical method. Using this LCMS approach, we characterized proteins isolated from E. coli grown in glucose, lactose, and acetate. The change in relative abundance of the corresponding proteins was measured from peptides common to both conditions. Protein identities were also determined for those peptides that were unique to each condition, and these identities were found to be consistent with the underlying biochemical restrictions imposed by the growth conditions. The relative change in abundance of the characterized proteins ranged from 0.1- to 90-fold among the three binary comparisons. The overall coverage of the characterized proteins ranged from 10 to 80%, consisting of one to 34 peptides per protein. The quantitative results obtained from our study were comparable to other existing proteomic and transcriptional profiling approaches. This study illustrates the robustness of this novel LCMS approach for the simultaneous quantitative and comprehensive qualitative analysis of proteins in complex mixtures.