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Plant N-glycan profiling of minute amounts of material

Martial Séveno 1, 2 Gleysin Cabrera 3 Ada Triguero 3 Carole Burel 1 Jérôme Leprince 4 Christophe Rihouey 5 Louis-Philippe Vezina 2 Marc-André d'Aoust 2 Pauline Rudd 6 L. Royle 7 Raymond Dwek 7 David Harvey 8 Patrice Lerouge 1 José Cremata 3 Muriel Bardor 1
1 GTCV - Glycobiologie et transports chez les végétaux
2 Medicago Inc.
3 CIGB - Center for Genetic Engineering and Biotechnology [Cuba]
4 Neuroendocrinologie cellulaire et moléculaire
5 PBS - Polymères Biopolymères Surfaces
6 UCD - University College Dublin [Dublin]
7 Oxford Glycobiology Institute
8 UNSW - School of Mathematics and Statistics [Sydney]
Abstract : Development of convenient strategies for identification of plant N-glycan profiles has been driven by the emergence of plants as an expression system for therapeutic proteins. In this article, we reinvestigated qualitative and quantitative aspects of plant N-glycan profiling. The extraction of plant proteins through a phenol/ammonium acetate procedure followed by deglycosylation with peptide N-glycosidase A (PNGase A) and coupling to 2-aminobenzamide provides an oligosaccharide preparation containing reduced amounts of contaminants from plant cell wall polysaccharides. Such a preparation was also suitable for accurate qualitative and quantitative evaluation of the N-glycan content by mass spectrometry. Combining these approaches allows the profiling to be carried out from as low as 500 mg of fresh leaf material. We also demonstrated that collision-induced dissociation (CID) mass spectrometry in negative mode of N-glycans harboring alpha(1,3)- or alpha(1,6)-fucose residue on the proximal GlcNAc leads to specific fragmentation patterns, thereby allowing the discrimination of plant N-glycans from those arising from mammalian contamination.
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https://hal-normandie-univ.archives-ouvertes.fr/hal-01974002
Contributor : Jérôme Leprince <>
Submitted on : Tuesday, January 8, 2019 - 3:19:02 PM
Last modification on : Thursday, July 2, 2020 - 3:30:07 AM

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COMUE-NORMANDIE | CNRS | INSA-ROUEN | UNIROUEN | INSA-GROUPE | PBS | ENSICAEN | UNILEHAVRE | UNICAEN

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Martial Séveno, Gleysin Cabrera, Ada Triguero, Carole Burel, Jérôme Leprince, et al.. Plant N-glycan profiling of minute amounts of material. Analytical Biochemistry, Elsevier Masson, 2008, 379 (1), pp.66-72. ⟨10.1016/j.ab.2008.04.034⟩. ⟨hal-01974002⟩

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