Mapping site‐specific protein N‐glycosylations through liquid chromatography/mass spectrometry and targeted tandem mass spectrometry |
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| Authors: | Yin Wu Yehia Mechref Iveta Klouckova Anoop Mayampurath Milos V. Novotny Haixu Tang |
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| Affiliation: | 1. School of Informatics and Computing, Indiana University, Bloomington, IN 47408, USA;2. Department of Chemistry, Indiana University, Bloomington, IN 47408, USA;3. National Center for Glycomics and Glycoproteomics, Indiana University, Bloomington, IN 47408, USA |
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| Abstract: | Glycosylation is one of the most common posttranslational modifications (PTMs) of proteins, the characterization of which is commonly achieved through proteomic protocol, involving trypsin digestion followed by liquid chromatography/tandem mass spectrometry (LC/MS/MS). However, it is often not possible to characterize all glycopeptides in a complex sample because of the high complexity of glycoproteomic samples, and the relative lower abundances of glycopeptides in comparison to the unmodified peptides. We present here a targeted MS/MS analysis approach, which utilizes a previously developed computational tool, GlyPID, to guide multiple experiments, thus permitting a complete characterization of all N‐glycosylation sites of glycoproteins present in a complex sample. We have tested our approach using model glycoproteins analyzed by high‐resolution LTQ‐FT MS. The results demonstrate a potential use of our method for a high‐throughput characterization of complex mixtures of glycosylated proteins. Copyright © 2010 John Wiley & Sons, Ltd. |
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