Ethier Martin, Saba Julian A, Ens Werner, Standing Kenneth G, Perreault Hélène
Department of Chemistry, University of Manitoba, Winnipeg, MB, Canada.
Rapid Commun Mass Spectrom. 2002;16(18):1743-54. doi: 10.1002/rcm.779.
Glycoprotein function is controlled by several biological factors, one of them being the structure of carbohydrate chains (glycans) attached to specific amino acids of the protein backbone. Changes in glycan structures have been shown to modify the secondary and tertiary conformation of glycoproteins, thus their function. Powerful analytical tools are available for the characterization of sugar structures, and recently mass spectrometry (MS) has been increasingly useful for this purpose. Manual interpretation of tandem mass spectrum is possible but tedious. Automated interpretation should speed the analysis and enhance the results obtained. A new computer program for automated interpretation of tandem MS spectra of complex N-linked glycans oligosaccharides from mammals will be described. N-Linked oligosaccharides standards were derivatized with 1-phenyl-3-methyl-5-pyrazolone (PMP) and analyzed by matrix-assisted laser desorption/ionization (MALDI)-tandem MS. Simulated tandem mass spectra of other common glycans were also generated to test the algorithm. The MALDI-MS/MS spectra featured resolved isotopic distributions for the M + H and fragment ions of oligosaccharides. These isotopic distributions complicated the automated analysis of the spectra and were removed to leave only monoisotopic peaks. An algorithm was written for this purpose, yielding simplified tandem mass spectra. Another algorithm is then used to determine the structure of the oligosaccharide. A score is then given to each structure, depending on agreement with experimental results. The program successfully assigned the true structure in 24 out of the 28 cases (86%) and the true structure was among the three top scoring structures in all cases.
糖蛋白的功能受多种生物学因素控制，其中之一是连接在蛋白质主链特定氨基酸上的碳水化合物链（聚糖）的结构。研究表明，聚糖结构的变化会改变糖蛋白的二级和三级构象，进而影响其功能。目前已有强大的分析工具用于糖结构的表征，近年来质谱（MS）在这方面的应用越来越广泛。手动解读串联质谱图是可行的，但很繁琐。自动解读应能加快分析速度并提高所得结果的质量。本文将介绍一种用于自动解读哺乳动物复杂N - 连接聚糖寡糖串联质谱图的新计算机程序。N - 连接寡糖标准品用1 - 苯基 - 3 - 甲基 - 5 - 吡唑啉酮（PMP）进行衍生化，并通过基质辅助激光解吸/电离（MALDI）- 串联质谱进行分析。还生成了其他常见聚糖的模拟串联质谱图以测试该算法。MALDI - MS/MS谱图显示了寡糖的M + H和碎片离子的分辨同位素分布。这些同位素分布使谱图的自动分析变得复杂，因此将其去除，仅保留单同位素峰。为此编写了一种算法，得到简化的串联质谱图。然后使用另一种算法确定寡糖的结构。接着根据与实验结果的吻合程度为每个结构打分。该程序在28个案例中的24个（86%）成功确定了真实结构，并且在所有案例中真实结构都在得分最高的三个结构之中。
