title: The binding sites of monoclonal antibodies to the non-reducing end of Francisella tularensis O-antigen accommodate mainly the terminal saccharide.
abstract: We have previously described two types of protective B-cell epitopes in the O-antigen (OAg) of the Gram-negative bacterium Francisella tularensis: repeating internal epitopes targeted by the vast majority of anti-OAg monoclonal antibodies (mAbs), and a non-overlapping epitope at the non-reducing end targeted by the previously unique IgG2a mAb FB11. We have now generated and characterized three mAbs specific for the non-reducing end of F. tularensis OAg, partially encoded by the same variable region germline genes, indicating that they target the same epitope. Like FB11, the new mAbs, Ab63 (IgG3), N213 (IgG3) and N62 (IgG2b), had higher antigen-binding bivalent avidity than internally binding anti-OAg mAbs, and an oligosaccharide containing a single OAg repeat was sufficient for optimal inhibition of their antigen-binding. The X-ray crystal structure of N62 Fab showed that the antigen-binding site is lined mainly by aromatic amino acids that form a small cavity, which can accommodate no more than one and a third sugar residues, indicating that N62 binds mainly to the terminal Qui4NFm residue at the nonreducing end of OAg. In efficacy studies with mice infected intranasally with the highly virulent F. tularensis strain SchuS4, N62, N213 and Ab63 prolonged survival and reduced blood bacterial burden. These results yield insights into how antibodies to non-reducing ends of microbial polysaccharides can contribute to immune protection despite the smaller size of their target epitopes compared with antibodies to internal polysaccharide regions.
authors.affiliation: Department of Pathology and Laboratory Medicine, Boston University School of Medicine, Boston, MA, USA.
authors.forename: Zhaohua
authors.initials: Z
authors.lastname: Lu
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Rynkiewicz
authors.forename: Chiou-Ying
authors.initials: CY
authors.lastname: Yang
authors.forename: Guillermo
authors.initials: G
authors.lastname: Madico
authors.forename: Hillary M
authors.initials: HM
authors.lastname: Perkins
authors.forename: Qi
authors.initials: Q
authors.lastname: Wang
authors.forename: Catherine E
authors.initials: CE
authors.lastname: Costello
authors.forename: Joseph
authors.initials: J
authors.lastname: Zaia
authors.forename: Barbara A
authors.initials: BA
authors.lastname: Seaton
authors.forename: Jacqueline
authors.initials: J
authors.lastname: Sharon
mesh_terms: D000818:Animals; D018033:Antibodies, Bispecific / Q000378:metabolism*; D000911:Antibodies, Monoclonal / Q000008:administration & dosage / Q000737:chemistry / Q000276:immunology*; D000915:Antibody Affinity; D058491:Bacterial Load; D001666:Binding Sites, Antibody; D002478:Cells, Cultured; D003460:Crystallization; D018985:Epitopes, B-Lymphocyte / Q000737:chemistry / Q000276:immunology / Q000378:metabolism*; D005260:Female; D005604:Francisella tularensis / Q000276:immunology*; D006801:Humans; D056724:Immunity, Humoral; D051379:Mice; D008807:Mice, Inbred BALB C; D019081:O Antigens / Q000276:immunology*; D011485:Protein Binding; D011487:Protein Conformation; D014406:Tularemia / Q000276:immunology* / Q000628:therapy
chemical_list: D018033:Antibodies, Bispecific; D000911:Antibodies, Monoclonal; D018985:Epitopes, B-Lymphocyte; D019081:O Antigens
keywords: bacteria/bacterial immunity; endotoxin/lipopolysaccharide; epitopes; immunoglobulins; vaccines