Proteomic Profiling of Intact Proteins Using WAX-RPLC 2-D Separations and FTICR Mass Spectrometry

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• 作者：Seema Sharma ; David C. Simpson ; Nikola Toli//pubs.acs.org/images/entities/cacute.gif" border="0"> ; Navdeep Jaitly ; Anoop M. Mayampurath ; Richard D. Smith ; Ljiljana Pa//pub
• 刊名：Journal of Proteome Research
• 出版年：2007
• 出版时间：February 2007
• 年：2007
• 卷：6
• 期：2
• 页码：602 - 610
• 全文大小：400K
• 年卷期：v.6,no.2(February 2007)
• ISSN：1535-3907

文摘

We investigated the combination of weak anion exchange (WAX) fractionation and on-line reversed-phase liquid chromatography (RPLC) separation using a 12 T FTICR mass spectrometer for the detectionof intact proteins from a Shewanella oneidensis MR-1 cell lysate. This work aimed at optimizing intactprotein detection for profiling proteins at a level that incorporates their modification state. A total of715 intact proteins were detected, and the combined results from the WAX fractions and theunfractionated cell lysate were aligned using LC-MS features to facilitate protein abundance measurements. Protein identifications and post-translational modifications were assigned for ~10% of thedetected proteins by comparing intact protein mass measurements to proteins identified in peptideMS/MS analysis of an aliquot of the same fraction. Intact proteins were also detected for S. oneidensislysates obtained from cells grown on ¹³C-, ¹⁵N-depleted media under aerobic and sub-oxic conditions.The strategy can be readily applied for measuring differential protein abundances and provides aplatform for high-throughput selection of biologically relevant targets for further characterization.