Springer Protocols: Abstract: Enzymatic Sequence Analysis of N-Glycans by Exoglycosidase Cleavage and Mass Spectrometry

Enzymatic Sequence Analysis of N-Glycans by Exoglycosidase Cleavage and Mass Spectrometry – detection of Lewis X Structures

By: Christoph Kannicht³, Detlef Grunow⁴, Lothar Lucka⁴

Abstract

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Enzymatic sequencing of oligosaccharides gives structural information on sequence of monosaccharides and type of linkage within the oligosaccharide chain. This data can be obtained by stepwise enzymatic digestion of a single, isolated oligosaccharide using individual or mixtures of specific exoglycosidases. N-glycans have to be fractionated from mixtures prior to sequence analysis to assign this type of structural information to a specific glycan. Enzymatic sequencing can be applied to oligosaccharide mixtures as well to evaluate the occurrence of distinct oligosac-charide motives of functional and/or structural interest.

Here we describe the application of enzymatic sequence analysis to a mixture of N-glycans released from α1-acid glycoprotein. The experimental conditions are optimized for detection of possible Lewis X structures after stepwise exoglycosi-dase digestion by MALDI-TOF mass spectrometry. However, the described method is generally applicable to analyze other structural properties of N-glycans by use of the respective specific exoglycosidases.

Affiliation(s): (3) Molecular Biochemistry Department, Octapharma Research and Development, Berlin, Germany
(4) Charite Universitätsmedizin Berlin, Institute for Biochemistry and Molecular Biology, Berlin, Germany

Book Title: Post-translational Modifi cations of Proteins: Tools for Functional Proteomics

Series: Methods in Molecular Biology | Volume: 446 | Pub. Date: Apr-04-2008 | Page Range: 255-266 | DOI: 10.1007/978-1-60327-084-7_18

Subject: Protein Science

Key Words: Enzymatic sequencing - N-glycan - exoglycosidase - MALDI-TOF - Lewis x - oligosaccharide structure

References

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