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Helm, J; Grunwald-Gruber, C; Thader, A; Urteil, J; Fuhrer, J; Stenitzer, D; Maresch, D; Neumann, L; Pabst, M; Altmann, F.
(2021): Bisecting Lewis X in Hybrid-Type N-Glycans of Human Brain Revealed by Deep Structural Glycomics
ANAL CHEM. 2021; 93(45): 15175-15182. FullText FullText_BOKU

The importance of protein glycosylation in the biomedical field requires methods that not only quantitate structures by their monosaccharide composition, but also resolve and identify the many isomers expressed by mammalian cells. The art of unambiguous identification of isomeric structures in complex mixtures, however, did not yet catch up with the fast pace of advance of high-throughput glycomics. Here, we present a strategy for deducing structures with the help of a deci-minute accurate retention time library for porous graphitic carbon chromatography with mass spectrometric detection. We implemented the concept for the fundamental N-glycan type consisting of five hexoses, four N-acetylhexosamines and one fucose residue. Nearly all of the 40 biosynthetized isomers occupied unique elution positions. This result demonstrates the unique isomer selectivity of porous graphitic carbon. With the help of a rather tightly spaced grid of isotope-labeled internal N-glycan, standard retention times were transposed to a standard chromatogram. Application of this approach to animal and human brain N-glycans immediately identified the majority of structures as being of the bisected type. Most notably, it exposed hybrid-type glycans with galactosylated and even Lewis X containing bisected N-acetylglucosamine, which have not yet been discovered in a natural source. Thus, the time grid approach implemented herein facilitated discovery of the still missing pieces of the N-glycome in our most noble organ and suggests conjunction with collision induced a starting point for the overdue development of isomer-specific deep structural glycomics.
Authors BOKU Wien:
Altmann Friedrich
Führer Johannes
Grünwald-Gruber Clemens
Helm Johannes
Maresch Daniel
Neumann Laura
Pabst Martin
Stenitzer David
Thader Andreas
Urteil Jonathan
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