N-linked oligosaccharide analysis of rat brain Thy-1 by liquid chromatography with graphitized carbon column/ion trap-Fourier transform ion cyclotron resonance mass spectrometry in positive and negative ion modes
N-linked oligosaccharide analysis of rat brain Thy-1 by liquid chromatography with graphitized carbon column/ion trap-Fourier transform ion cyclotron resonance mass spectrometry in positive and negative ion modes
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DOI：
10.1016/j.chroma.2005.11.043
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发表时间：
2006-01-27
影响因子：
4.1
通讯作者：
Kawanishi, T
中科院分区：
文献类型：
--
作者：
Itoh, S;Kawasaki, N;Kawanishi, T
我们先前已经描述了使用蛋白酶消化的Thy-1通过LC/多级串联质谱（MS '）对大鼠脑Thy-1的位点特异性糖基化分析。在本研究中，通过使用石墨化碳柱的LC/MS（GCC-LC/MS）的质谱寡糖分析来阐明从Thy-1释放的寡糖的详细结构。首先，使用模型寡糖，我们改进了寡糖的离子阱质谱（IT-NIS）与傅立叶变换离子回旋共振质谱（FT-ICR-MS）联用。采用FT-ICR-MS进行全MSn扫描，然后采用正离子模式的IT-MS进行数据依赖性MS'，以及随后采用FT-ICR-MS进行全MS 1扫描，然后采用负离子模式的IT-MS进行数据依赖性MSn，这一顺序扫描使得能够在单次分析中进行单糖组成分析以及中性和酸性寡糖的分析和测序。应用改进的寡糖谱法对十二烷基硫酸钠-聚丙烯酰胺凝胶电泳分离的Thy-1中的N-连接寡糖进行了分析。结果表明，Thy-1具有多种N-连接寡糖，包括刘易斯a/x、刘易斯B/y，部分结构为双唾液酸化结构。该方法适用于少量糖蛋白的分析，有望成为糖蛋白组学研究的有力工具。(c)2005 Elsevier B. V.保留所有权利。
We have previously described the site-specific glycosylation analysis of rat brain Thy-1 by LC/multistage tandem mass spectrometry (MS') using proteinase-digested Thy-1. In the present study, detailed structures of oligosaccharides released from Thy-1 were elucidated by mass spectrometric oligosaccharide profiling using LC/MS with a graphitized carbon column (GCC-LC/MS). First, using model oligosaccharides, we improved the oligosaccharide profiling by ion trap mass spectrometry (IT-NIS) coupled with Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR-MS). Sequential scanning of a full MSn scan with FT-ICR-MS followed by data-dependent MS' with IT-MS in positive ion mode, and a subsequent full MS1 scan with FT-ICR-MS followed by data-dependent MSn with IT-MS in negative ion mode enabled the monosaccharide composition analysis as well as profiling and sequencing of both neutral and acidic oligosaccharides in a single analysis. The improved oligosaccharide profiling was applied to elucidation of N-linked oligosaccharides from Thy-1 isolated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. It was demonstrated that Thy-1 possesses a significant variety of N-linked oligosaccharides, including Lewis a/x, Lewis b/y, and disialylated structure as a partial structure. Our method could be applicable to analysis of a small abundance of glycoproteins, and could become a powerful tool for glycoproteomics. (c) 2005 Elsevier B.V. All rights reserved.