EXPLORATIONS OF CAPILLARY ELECTROPHORESIS IN THE ANALYSIS OF MODEL GLYOPROTEINS

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 近年来，糖基化对蛋白质的作用受到越来越多的研究，因为所产生的糖蛋白已被证明影响蛋白质的功能和活性。因为可以存在多个糖基化位点，并且可能与每个位点关联大量的糖链结构，所以可能的结构组合的数量通常非常大。结合的可能数量随着蛋白质分子大小的增加而大大增加。事实证明，葡聚糖结构和各种连接点的解析和鉴定是一项极具挑战性的分析项目，尚未得到充分解决。在糖链分析中，即使是最强大的质谱学技术也不能对等压结构进行频率解析。我们建议使用一组模型糖蛋白来研究毛细管电泳法在糖蛋白分析中的潜力。首先，我们将利用毛细管电泳联用激光诱导荧光检测技术来检测模型糖蛋白胰酶消化过程中糖肽的硼酸盐络合引起的凝胶迁移率变化的可能性。这项技术可以快速识别哪些多肽是糖基化的。我们还将探索毛细管电泳质谱在分析复杂的糖蛋白混合物方面的潜力，这些混合物是用基于硼酸盐的电解液从模型糖蛋白中酶解出来的。这两种相关的分析技术都可以为医学上有重要意义的糖蛋白的结构分析提供关键信息，并可能发展成为常规糖组分分析的强大工具。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. In recent years the role of glycosylation to proteins has been under increased investigation as the resulting glycoproteins have been shown to influence protein function and activity. Because multiple glycosylation sites can exist and a large number of glycan structures can potentially be associated with each site, the number of possible structural combinations is frequently very large. The possible number of combination increases greatly with the size of the molecular size of the protein. The resolution and identification of glycan structure and the various points of attachment has proven to be very challenging analytical project and has not been adequately addressed. Even the most powerful mass spectrometric techniques cannot frequency resolve isobaric structures in glycan analysis. We propose to investigate the potential of capillary electrophoresis in the analysis of glycoproteins using a group of model glycoproteins. First, we will examine the potential of electrophoretic mobility shifts induced by borate complexation of glycopeptides in trypsin digest of model glycoproteins using capillary electrophoresis with laser induced fluorescence detection. This technique could rapidly identify which peptides are glycosylated. We will also explore the potential of capillary electrophoresis-mass spectrometry in the analysis of complex glycan mix1ures that have been enzymatically released from model glycoproteins using borate-based electrolytes. Both of these related analysis techniques could provide critical information in the structural analysis of medically significant glycoproteins and could develop into powerful tools in routine glycomics analysis.