High-Speed Data Reduction, Feature Detection, and MS/MS Spectrum Quality Assessment of Shotgun Proteomics Data Sets Using High-Resolution Mass Spectrometry

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• 作者：Michael R. Hoopmann ; Gregory L. Finney ; Michael J. MacCoss
• 刊名：Analytical Chemistry
• 出版年：2007
• 出版时间：August 1, 2007
• 年：2007
• 卷：79
• 期：15
• 页码：5620 - 5632
• 全文大小：644K
• 年卷期：v.79,no.15(August 1, 2007)
• ISSN：1520-6882

文摘

Advances in Fourier transform mass spectrometry havemade the acquisition of high-resolution and accurate massmeasurements routine on a chromatographic time scale.Here we report an algorithm, Hardklör, for the rapid androbust analysis of high-resolution mass spectra acquiredin shotgun proteomics experiments. Our algorithm isdemonstrated in the analysis of an Escherichia colienriched membrane fraction. The mass spectrometry dataof the respective peptides are acquired by microcapillaryHPLC on an LTQ-orbitrap mass spectrometer with data-dependent acquisition of MS/MS spectra. Hardklör detects 211,272 total peptide isotope distributions over a2-h analysis (75-min gradient) in only a small fraction ofthe time required to acquire the data. From these datathere are 13 665 distinct, chromatographically persistentpeptide isotope distributions. Hardklör is also used toassess the quality of the product ion spectra and finds thatmore than 11.2% of the MS/MS spectra are composed offragment ions from multiple different molecular species.Additionally, a method is reported that enzymaticallylabels N-linked glycosylation sites on proteins, creating aunique isotope signature that can be detected with Hardklör. Using the protein invertase, Hardklör identifies ¹⁸O-labeled peptide isotope distributions of four glycosylationsites. The speed and robustness of the algorithm create aversatile tool that can be used in many different areas ofmass spectrometry data analysis.