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Gewählte Publikation:

Yan, S; Vanbeselaere, J; Jin, CS; Blaukopf, M; Wols, F; Wilson, LBH; Paschinger, K.
(2018): Core Richness of N-Glycans of Caenorhabditis elegans: A Case Study on Chemical and Enzymatic Release
ANAL CHEM. 2018; 90(1): 928-935. FullText FullText_BOKU

Despite years of research, the glycome of the model nematode Caenorhabditis elegans is still not fully understood. Certainly, data over the years have indicated that this organism synthesizes unusual N-glycans with a range of galactose and fucose modifications on the Man(2-3)GlcNAc(2), core region. Previously, up to four fucose residues were detected on its N-glycans, despite these lacking the fucosylated antennae typical of many other eukaryotes; some of these fucose residues are capped with hexose residues as shown by the studies of us and others. There have, though, been contrasting reports regarding the maximal number of fucose substitutions in C. elegans, which in part may be due to different methodological approaches, including use of either peptide:N-glycosidases F and A (PNGase F and A) or anhydrous hydrazine to cleave the N-glycans from glycopeptides. Here we compare the use of hydrazine with that of a new enzyme (rice PNGase Ar) and show that both enable release of glycans with more sugar residues on the proximal GlcNAc than previously resolved. By use of exoglycosidase sequencing, in conjunction with high-performance liquid chromatography (HPLC) and matrix-assisted laser desorption ionization time-of-flight tandem mass spectrometry (MALDI-TOF MS/MS), we now reveal that actually up to five fucose residues modify the core region of C. elegans N-glycans and that the alpha 1,3-fucose on the reducing terminus can be substituted by an a-linked galactose. Thus, traditional PNGase F and A release may be insufficient for release of the more highly core-modified N-glycans, especially those occurring in C. elegans, but novel enzymes can compete against chemical methods in terms of safety, ease of cleanup, and quality of resulting glycomic data.
Autor/innen der BOKU Wien:
Blaukopf Markus
Paschinger Katharina
Vanbeselaere Jorick
Wilson Iain B.H.
Yan Shi
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