EXPLORATIONS OF CAPILLARY ELECTROPHORESIS IN THE ANALYSIS OF MODEL GLYOPROTEINS

该子项目是利用 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得主要资金， 因此可以在其他CRISP条目中表示。列出的机构是 中心，不一定是研究者的机构。 近年来，糖基化对蛋白质的作用受到越来越多的研究，因为所得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，并且大量聚糖结构可以潜在地与每个位点相关联，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。聚糖结构和各种连接点的解析和鉴别已被证明是非常具有挑战性的分析项目，尚未得到充分解决。即使是最强大的质谱技术也不能在聚糖分析中对同量异位素结构进行频率分辨。我们建议使用一组模型糖蛋白来研究毛细管电泳在糖蛋白分析中的潜力。首先，我们将研究由硼酸盐络合的糖肽在胰蛋白酶消化的模型糖蛋白，使用毛细管电泳与激光诱导荧光检测诱导的电泳迁移率变化的潜力。该技术可以快速鉴定哪些肽是糖基化的。我们还将探讨毛细管电泳-质谱法在分析复杂的聚糖混合物中的潜力，这些聚糖混合物是使用硼酸盐基电解质从模型糖蛋白中酶促释放的。这两种相关的分析技术可以提供重要的信息，在医学上重要的糖蛋白的结构分析，并可能发展成为常规糖组学分析的强大工具。

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.