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.
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