Structural characterization of glycoproteins using ms

University Of Kansas Lawrence, Lawrence KS

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Abstract

The glycoprotein hormones to be studied are used as diagnostic markers, for pregnancy and tumors, and as pharmaceuticals, for the treatment of infertility and thyroid cancer. Thus, a reliable method to characterize these proteins, particularly when they are used for medicinal purposes, is paramount in order to assure medication is safe and effective. Current techniques used to study these compounds are time-consuming. Furthermore, they often do not provide sufficient information about the point of attachment, of the carbohydrate to the protein, or sufficient information about the heterogeneity of the carbohydrates. We propose to overcome these problems by analyzing intact glycopeptides using tandem mass spectrometry. By developing mass spectrometric methods of characterizing the carbohydrates, while they remain attached to the protein, the origin of carbohydrate attachment will be unambiguous. Our approach will utilize tools developed for analyzing carbohydrates that are not linked to proteins; however we will tailor these techniques to accommodate the peptide fragments linked to the reducing end of the oligosaccharides. After demonstrating our technique will be successful on a purified glycopeptide, we will develop the appropriate separation strategies required to use our technology in characterizing a glycoprotein that has multiple glycosylation sites.