title: 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.
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)ABAc CAc D]) repeating unit, and the non-O-acetylated equivalent defines S. flexneri X. Propyl hepta-, octa-, and decasaccharides sharing the (E')A'BAc CD(E)A sequence, and their non-O-acetylated analogues were synthesized from a fully protected BAc CD(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 BAc CD 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.
authors.affiliation: Institut Pasteur, Unité de Chimie des Biomolécules, 28 rue du Dr. Roux, 75724, Paris Cedex 15, France.|CNRS UMR 3523, Institut Pasteur, 75015, Paris, France.|Université Paris Descartes, Institut Pasteur, 75015, Paris, France.|Present address for J.B.: Adisseo (France), Present address for P.B., Dept. of Organic Chemistry, Arrhenius Laboratory, Stockholm University, 10691, Stockholm, Sweden.
authors.forename: Julien
authors.initials: J
authors.lastname: Boutet
authors.affiliation: Chemical and Physical Biology, Centro de Investigaciones Biológicas, CSIC, Ramiro de Maeztu 9, 28040, Madrid, Spain.|Present address for J.B.: Adisseo (France), Present address for P.B., Dept. of Organic Chemistry, Arrhenius Laboratory, Stockholm University, 10691, Stockholm, Sweden.
authors.forename: Pilar
authors.initials: P
authors.lastname: Blasco
authors.affiliation: Institut Pasteur, Unité de Chimie des Biomolécules, 28 rue du Dr. Roux, 75724, Paris Cedex 15, France.|CNRS UMR 3523, Institut Pasteur, 75015, Paris, France.
authors.forename: Catherine
authors.initials: C
authors.lastname: Guerreiro
authors.affiliation: Institut Pasteur, Unité de Pathogénie Microbienne Moléculaire, 28 rue du Dr. Roux, 75015, Paris, France.|INSERM U1202, Institut Pasteur, 75015, Paris, France.
authors.forename: Françoise
authors.initials: F
authors.lastname: Thouron
authors.affiliation: Institut Pasteur, PF5, 28 rue du Dr. Roux, 75015, Paris, France.|CNRS UMR 3528, Institut Pasteur, 75015, Paris, France.
authors.forename: Sylvie
authors.initials: S
authors.lastname: Dartevelle
authors.affiliation: Institut Pasteur, PF5, 28 rue du Dr. Roux, 75015, Paris, France.|CNRS UMR 3528, Institut Pasteur, 75015, Paris, France.
authors.forename: Farida
authors.initials: F
authors.lastname: Nato
authors.affiliation: Chemical and Physical Biology, Centro de Investigaciones Biológicas, CSIC, Ramiro de Maeztu 9, 28040, Madrid, Spain.
authors.forename: F Javier
authors.initials: FJ
authors.lastname: Cañada
authors.affiliation: Chemical and Physical Biology, Centro de Investigaciones Biológicas, CSIC, Ramiro de Maeztu 9, 28040, Madrid, Spain.|Molecular Recognition & Host-Pathogen Interactions Program, CIC bioGUNE, Bizkaia Technological Park, Building 801A, 48160, Derio, Spain.
authors.forename: Ana
authors.initials: A
authors.lastname: Ardá
authors.affiliation: Institut Pasteur, Unité de Pathogénie Microbienne Moléculaire, 28 rue du Dr. Roux, 75015, Paris, France.|INSERM U1202, Institut Pasteur, 75015, Paris, France.
authors.forename: Armelle
authors.initials: A
authors.lastname: Phalipon
authors.affiliation: Chemical and Physical Biology, Centro de Investigaciones Biológicas, CSIC, Ramiro de Maeztu 9, 28040, Madrid, Spain. jjbarbero@cicbiogune.es.|Molecular Recognition & Host-Pathogen Interactions Program, CIC bioGUNE, Bizkaia Technological Park, Building 801A, 48160, Derio, Spain. jjbarbero@cicbiogune.es.|Ikerbasque, Basque Foundation for Science, Maria Lopez de Haro 3, 48013, Bilbao, Spain. jjbarbero@cicbiogune.es.
authors.forename: Jesús
authors.initials: J
authors.lastname: Jiménez-Barbero
authors.affiliation: Institut Pasteur, Unité de Chimie des Biomolécules, 28 rue du Dr. Roux, 75724, Paris Cedex 15, France. laurence.mulard@pasteur.fr.|CNRS UMR 3523, Institut Pasteur, 75015, Paris, France. laurence.mulard@pasteur.fr.
authors.forename: Laurence A
authors.initials: LA
authors.lastname: Mulard
mesh_terms: D000818:Animals; D007120:Immunochemistry; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D051379:Mice; D008807:Mice, Inbred BALB C; D019081:O Antigens / Q000737:chemistry*; D012763:Shigella flexneri / Q000737:chemistry*
chemical_list: D019081:O Antigens
keywords: NMR spectroscopy; antibodies; carbohydrates; epitope specificity; glycosylation