MRM screening/biomarker discovery with linear ion trap MS: a library of human cancer-specific peptides.
MRM screening/biomarker discovery with linear ion trap MS: a library of human cancer-specific peptides.
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DOI：
10.1186/1471-2407-9-96
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发表时间：
2009-03-27
期刊：
影响因子：
3.8
通讯作者：
Lazar IM
中科院分区：
文献类型：
--
作者：
Yang X;Lazar IM
新的蛋白质生物标志物的发现在临床环境中是必不可少的，可以实现早期疾病诊断和提高生存率。为了促进差异表达分析和生物标志物的发现，各种基于串联质谱（MS/MS）的蛋白质分析技术已经被开发出来。为了实现灵敏的检测和准确的定量，已经实施了靶向质谱筛选方法，如多反应监测（MRM）。采用2d强阳离子交换(SCX)/反相液相色谱（RPLC）分离与线性离子阱质谱（MS）检测相结合的方法对MCF-7乳腺癌蛋白细胞提取物进行了分析。质谱数据用Sequest-based Bioworks软件（Thermo Electron）进行分析。使用内部开发的perl脚本计算光谱计数和每个肽的代表性片段离子。在这项工作中，我们报告了9,677个肽库的生成（p < 0.001），代表了来自人类乳腺癌细胞的约1,572个蛋白质，可用于MRM/MS-based生物标志物筛选研究。对于每种蛋白质，该文库提供可检测肽的数量和序列，电荷状态，光谱计数，分子量，表征串联质谱质量的参数(p值，DeltaM, Xcorr, DeltaCn, Sp, no。a, b， y离子在光谱中的匹配度)，保留时间，以及对应于给定肽的前10个最强烈的产物离子。只有被至少两个光谱计数识别的蛋白质被列出。蛋白质频率的实验分布，作为分子量的函数，与SwissProt数据库中提供的人类蛋白质组中蛋白质的理论分布密切匹配。所鉴定蛋白质的氨基酸序列覆盖率为0.04% ~ 98.3%。细胞提取物中丰度最高的蛋白质分子量(MW)<50,000。初步实验表明，在复杂的未分离样品中，传统的依赖数据的质谱采集方法无法检测到的假定的生物标志物，可以通过该文库提供的信息进行可靠的鉴定。根据谱计数、串联质谱的质量和母肽及其最丰富的子离子的m/z值，可以选择MRM条件来检测目标肽和蛋白质。
The discovery of novel protein biomarkers is essential in the clinical setting to enable early disease diagnosis and increase survivability rates. To facilitate differential expression analysis and biomarker discovery, a variety of tandem mass spectrometry (MS/MS)-based protein profiling techniques have been developed. For achieving sensitive detection and accurate quantitation, targeted MS screening approaches, such as multiple reaction monitoring (MRM), have been implemented. MCF-7 breast cancer protein cellular extracts were analyzed by 2D-strong cation exchange (SCX)/reversed phase liquid chromatography (RPLC) separations interfaced to linear ion trap MS detection. MS data were interpreted with the Sequest-based Bioworks software (Thermo Electron). In-house developed Perl-scripts were used to calculate the spectral counts and the representative fragment ions for each peptide. In this work, we report on the generation of a library of 9,677 peptides (p < 0.001), representing ~1,572 proteins from human breast cancer cells, that can be used for MRM/MS-based biomarker screening studies. For each protein, the library provides the number and sequence of detectable peptides, the charge state, the spectral count, the molecular weight, the parameters that characterize the quality of the tandem mass spectrum (p-value, DeltaM, Xcorr, DeltaCn, Sp, no. of matching a, b, y ions in the spectrum), the retention time, and the top 10 most intense product ions that correspond to a given peptide. Only proteins identified by at least two spectral counts are listed. The experimental distribution of protein frequencies, as a function of molecular weight, closely matched the theoretical distribution of proteins in the human proteome, as provided in the SwissProt database. The amino acid sequence coverage of the identified proteins ranged from 0.04% to 98.3%. The highest-abundance proteins in the cellular extract had a molecular weight (MW)<50,000. Preliminary experiments have demonstrated that putative biomarkers, that are not detectable by conventional data dependent MS acquisition methods in complex un-fractionated samples, can be reliable identified with the information provided in this library. Based on the spectral count, the quality of a tandem mass spectrum and the m/z values for a parent peptide and its most abundant daughter ions, MRM conditions can be selected to enable the detection of target peptides and proteins.
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影响因子：
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