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Universal Proteolysis and MSn for N- and O- Glycan Branching Analysis



John E. Schiel, Karen W. Phinney, Nicholas J. Smith


The continually growing list of critical glycosylation-related processes has made analytical methodology for detailed glycan characterization an area of increasing interest. Glycosylation is a post translational modification of unsurpassed complexity due to the variety of compositions and linkages formed by these biopolymers. Structural characterization of glycan isomers has been achieved using ion trap mass spectrometry and MSn of released, permethylated glycans. In some cases, however, release of the glycans may be hindered, result in degradation of the glycan, and produce limited yields of permethylated product. In the current report, we demonstrate universal proteolysis of both N- and O-linked glycoproteins to individual glycoamino acids. These samples were shown to be directly amenable to permethylation and MSn analysis for isomeric structural determination. Universal proteolysis and permethylation provides an identical sample preparation strategy for both classes of glycans that avoids potential pitfalls of commonly used release methods. This methodology should be applicable to all glycoproteins, and serve as an alternative to glycan release for MSn branching analysis.
Journal of Mass Spectrometry


Glycoprotein, Glycan, Mass Spectrometry, Permethylation, Branching


Schiel, J. , Phinney, K. and Smith, N. (2013), Universal Proteolysis and MSn for N- and O- Glycan Branching Analysis, Journal of Mass Spectrometry (Accessed June 20, 2024)


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Created April 20, 2013, Updated December 18, 2019