FOURIER TRANSFORM MASS SPECTROMETRY OF OLIGOSACCHARIDES

低聚糖异质性的分析将通过 基于傅立叶变换的几种互补分析方法的发展 变换质谱学。低聚糖与宿主有关 包括细胞-细胞和细胞-基质识别的生物功能， 荷尔蒙的作用、细胞间和细胞内的运输以及保护。 然而，与DNA和蛋白质不同的是，序列提供了几乎所有的 一级结构，低聚糖通过它们的序列来表征， 联系，和立体化学。此外，物种的巨大多样性 单糖由于化学修饰和异构化，不稳定 糖苷键的性质，以及较差的固有碱性 并对低聚糖进行结构解析 这比其他生物聚合物要难得多。目前有 对于具有敏感性的低聚糖，没有类似的方法， 蛋白质埃德曼降解的可靠性和准确性。我们 建议开发快速阐明结构的分析方法 在寡糖库中。这些方法将在 研究南非蟾蜍非洲爪哇的水母皮毛。X. Levis是早期研究的重要模型。 受精。果冻大衣在几个重要的 包括阻断多精受精、防止杂交等过程 施肥和保护。碰撞引起的解离将是 用于确定所释放的低聚糖的结构 阿迪托斯。结构基序目录及其对应的CID 将产生碎片图案(低聚糖指纹) 利用已知的低聚糖的结构。从这个目录中， 次要组件的结构将被确定。碱性 将使用退化(AD)来获得链接和序列信息 未知的低聚糖。AD与基质辅助激光器的耦合 解吸/电离和傅里叶变换质谱学 最近在该实验室中演示了提供序列和 关于一系列模型化合物的连接信息。该方法将是 进一步改进以获得更高的灵敏度和更短的分析时间。它 还将与电喷雾电离相结合。此方法将是 用莱维氏X.laevis的低聚糖进行测试。然而，它的应用 是针对结构信息很少的图书馆。 最后，将开发一种方法来消除繁琐的过程 在分析前进行分离。生物素-亲和素的强相互作用将是 为开发分析物特异性MALDI探针奠定了基础。一个 固定化亲和素探针将用于提取生物素化 直接从溶液中提取低聚糖用于即时MALDI-FTMS 分析。这种方法省去了必要的长时间分离过程 用于分离释放的低聚糖。

The analysis of oligosaccharide heterogeneity will be addressed by the development of several complementary analytical methods based on Fourier transform mass spectrometry. Oligosaccharides are involved in a host of biological functions including cell-cell and cell-matrix recognition, hormonal actions, inter- and intracellular trafficking, and protection. However, unlike DNA and proteins where sequence provides nearly all the primary structure, oligosaccharides are characterized by their sequence, linkage, and stereochemistry. Additionally, the large diversity in the monosaccharides due to chemical modification and isomerism, the labile nature of the glycosidic bonds, and the poor intrinsic basicity all combine to make the structural elucidation of oligosaccharides significantly more difficult than other biopolymers. There is currently no analogous method for oligosaccharides with the sensitivity, reliability and accuracy of the Edman degradation for proteins. We propose to develop analytical methods to rapidly elucidate structures in oligosaccharide libraries. The methods will be developed in the study of the jelly coat of the South African toad, Xenopus laevis. X. laevis is an important model for the study of the early stages of fertilization. The jelly coat plays a role in several important processes including the block to polyspermy, prevention of cross fertilization and protection. Collision induced dissociation will be employed to determine structures of the released oligosaccharide alditols. A catalog of structural motifs with their corresponding CID fragmentation pattern (an oligosaccharide fingerprint) will be produced using the structures of known oligosaccharides. From this catalog, the structure of the minor components will be determined. Alkaline degradation (AD) will be used to obtain linkage and sequence information of unknown oligosaccharides. AD coupled with matrix-assisted laser desorption/ionization and Fourier transform mass spectrometry has recently been demonstrated in this laboratory to provide sequence and linkage information on a host of model compounds. The method will be further refined for greater sensitivity and shorter analysis time. It will also be coupled with electrospray ionization. This method will be tested with the oligosaccharides of X. laevis. However, its application is aimed towards libraries where little structural information exists. Finally, a method will be developed to eliminate the tedious process of separation prior to analysis. Strong biotin-avidin interaction will be the basis for the development of an analyte specific MALDI probe. A probe with immobilized avidin will be used to extract biotinylated oligosaccharides directly from solution for immediate MALDI-FTMS analysis. This method bypasses the long separation process necessary for the isolation of released oligosaccharides.