Comparative Profiling of Serum Glycoproteome by Sequential Purification of Glycoproteins and 2-Nitrobenzensulfenyl (NBS) Stable Isotope Labeling: A New Approach for the Novel Biomarker Discovery for C

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• 作者：Koji Ueda ; Toyomasa Katagiri ; Takashi Shimada ; Shinji Irie ; Taka-Aki Sato ; Yusuke Nakamura ; Yataro Daigo
• 刊名：Journal of Proteome Research
• 出版年：2007
• 出版时间：September 2007
• 年：2007
• 卷：6
• 期：9
• 页码：3475 - 3483
• 全文大小：386K
• 年卷期：v.6,no.9(September 2007)
• ISSN：1535-3907

文摘

The recent progress in various proteomic technologies allows us to screen serum biomarker includingcarbohydrate antigens. However, only a limited number of proteins could be detected by currentconventional methods such as shotgun proteomics, primarily because of the enormous concentrationdistribution of serum proteins and peptides. To circumvent this difficulty and isolate potential cancer-specific biomarkers for diagnosis and treatment, we established a new screening system consisting ofthe sequential steps of (1) immunodepletion of 6 high-abundance proteins, (2) targeted enrichment ofglycoproteins by lectin column chromatography, and (3) the quantitative proteome analysis using ¹²C₆-or ¹³C₆-NBS (2-nitrobenzenesulfenyl) stable isotope labeling followed by MALDI-QIT-TOF massspectrometric analysis. Through this systematic analysis for five serum samples derived from patientswith lung adenocarcinoma, we identified as candidate biomarkers 34 serum glycoproteins that revealedsignificant difference in 1,6-fucosylation level between lung cancer and healthy control, clearlydemonstrating that the carbohydrate-focused proteomics could allow for the detection of serumcomponents with cancer-specific features. In addition, we developed a more simplified and practicaltechnique, mass spectrometry-based glycan structure analysis and lectin blotting, in order to validateglycan structure of candidate biomarkers that could be applicable in clinical use. Our new glycoproteomicstrategy will provide highly sensitive and quantitative profiling of specific glycan structures on multipleproteins, which should be useful for serum biomarker discovery.