Detailed Investigation of the Immunodominant Role of O‐Antigen Stoichiometric O‐Acetylation as Revealed by Chemical Synthesis,Immunochemistry, Solution Conformation and STD‐NMR Spectroscopy for Shigella flexneri 3a
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| Authors: | Dr Julien Boutet Dr Pilar Blasco Catherine Guerreiro Françoise Thouron Sylvie Dartevelle Dr Farida Nato Dr F Javier Cañada Dr Ana Ardá Dr Armelle Phalipon Dr Jesús Jiménez‐Barbero Dr Laurence A Mulard |
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| Affiliation: | 1. Institut Pasteur, Unité de Chimie des Biomolécules, Paris Cedex 15, France;2. CNRS UMR 3523, Institut Pasteur, Paris, France;3. Université Paris Descartes, Institut Pasteur, Paris, France;4. Present address for J.B.: Adisseo (France), Present address for P.B., Dept. of Organic Chemistry, Arrhenius Laboratory, Stockholm University, Stockholm, Sweden;5. Chemical and Physical Biology, Centro de Investigaciones Biológicas, CSIC, Madrid, Spain;6. Institut Pasteur, Unité de Pathogénie Microbienne Moléculaire, Paris, France;7. INSERM U1202, Institut Pasteur, Paris, France;8. Institut Pasteur, PF5, Paris, France;9. CNRS UMR 3528, Institut Pasteur, 75015, Paris, France;10. Molecular Recognition & Host-Pathogen Interactions Program, CIC bioGUNE, Bizkaia Technological Park, Building 801A, Derio, Spain;11. Ikerbasque, Basque Foundation for Science, Bilbao, Spain |
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| Abstract: | Shigella flexneri 3a causes bacillary dysentery. Its O‐antigen has the {2)‐α‐d ‐Glcp‐(1→3)]‐α‐l ‐Rhap‐(1→2)‐α‐l ‐Rhap‐(1→3)‐Ac→2]‐α‐l ‐Rhap‐(1→3)‐Ac→6]≈40 %‐β‐d ‐GlcpNAc‐(1→} ((E)ABAcCAcD]) repeating unit, and the non‐O‐acetylated equivalent defines S. flexneri X. Propyl hepta‐, octa‐, and decasaccharides sharing the (E′)A′BAcCD(E)A sequence, and their non‐O‐acetylated analogues were synthesized from a fully protected BAcCD(E)A allyl glycoside. The stepwise introduction of orthogonally protected mono‐ and disaccharide imidate donors was followed by a two‐step deprotection process. Monoclonal antibody binding to twenty‐six S. flexneri types 3a and X di‐ to decasaccharides was studied by an inhibition enzyme‐linked immunosorbent assay (ELISA) and STD‐NMR spectroscopy. Epitope mapping revealed that the 2C‐acetate dominated the recognition by monoclonal IgG and IgM antibodies and that the BAcCD segment was essential for binding. The glucosyl side chain contributed to a lesser extent, albeit increasingly with the chain length. Moreover, tr‐NOESY analysis also showed interaction but did not reveal any meaningful conformational change upon antibody binding. |
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| Keywords: | antibodies carbohydrates epitope specificity glycosylation NMR spectroscopy |
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