Kubota Kazutosi, Sato Yuji, Suzuki Yusuke, Goto-Inoue Naoko, Toda Tosifusa, Suzuki Minoru, Hisanaga Shin-ichi, Suzuki Akemi, Endo Tamao
Research Team for Functional Genomics and Research Team for Molecular Biomarkers, Tokyo Metropolitan Institute of Gerontology, Foundation for Research on Aging and Promotion of Human Welfare, 35-2 Sakae-cho, Itabashi-ku, Tokyo 173-0015, Japan.
Anal Chem. 2008 May 15;80(10):3693-8. doi: 10.1021/ac800070d. Epub 2008 Apr 15.
Glycopeptides prepared from 1 nmol of a mixture of glycoproteins, transferrin, and ribonuclease B by lysylendopeptidase digestion were isolated by lectin and cellulose column chromatographies, and then they were analyzed by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry and MALDI-quadrupole ion trap (QIT)-TOF mass spectrometry which enables the performance of MS ( n ) analysis. The lectin affinity preparation of glycopeptides with Sambucus nigra agglutinin and concanavalin A provides the glycan structure outlines for the sialyl linkage and the core structure of N-glycans. Such structural estimation was confirmed by MALDI-TOF MS and MALDI-QIT-TOF MS/MS. Amino acid sequences and location of glycosylation sites were determined by MALDI-QIT-TOF MS/MS/MS. Taken together, the combination of lectin column chromatography, MALDI-TOF MS, and MALDI-QIT-TOF MS ( n ) provides an easy way for the structural estimation of glycans and the rapid analysis of glycoproteomics.
通过赖氨酰内肽酶消化从1纳摩尔糖蛋白、转铁蛋白和核糖核酸酶B的混合物中制备的糖肽，经凝集素和纤维素柱色谱分离，然后用基质辅助激光解吸/电离飞行时间（MALDI-TOF）质谱和MALDI-四极杆离子阱（QIT）-TOF质谱进行分析，后者能够进行MS（n）分析。用黑接骨木凝集素和伴刀豆球蛋白A对糖肽进行凝集素亲和制备，可提供唾液酸连接和N-聚糖核心结构的聚糖结构轮廓。这种结构估计通过MALDI-TOF MS和MALDI-QIT-TOF MS/MS得到证实。通过MALDI-QIT-TOF MS/MS/MS确定氨基酸序列和糖基化位点的位置。综上所述，凝集素柱色谱、MALDI-TOF MS和MALDI-QIT-TOF MS（n）的结合为聚糖的结构估计和糖蛋白质组学的快速分析提供了一种简便方法。
