The binding sites of monoclonal antibodies to the non‐reducing end of F rancisella tularensis O ‐antigen accommodate mainly the terminal saccharide

We have previously described two types of protective B ‐cell epitopes in the O ‐antigen ( OA g) of the Gram‐negative bacterium F rancisella tularensis : repeating internal epitopes targeted by the vast majority of anti‐ OA g monoclonal antibodies ( mA bs), and a non‐overlapping epitope at the non‐reducing end targeted by the previously unique IgG2a mA b FB 11. We have now generated and characterized three mA bs specific for the non‐reducing end of F . tularensis OA g, partially encoded by the same variable region germline genes, indicating that they target the same epitope. Like FB11, the new mA bs, A b63 (Ig G 3), N213 (Ig G 3) and N 62 (Ig G 2b), had higher antigen‐binding bivalent avidity than internally binding anti‐ OA g mA bs, and an oligosaccharide containing a single OA g repeat was sufficient for optimal inhibition of their antigen‐binding. The X ‐ray crystal structure of N 62 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 Qui4 NF m residue at the nonreducing end of OA g. In efficacy studies with mice infected intranasally with the highly virulent F . tularensis strain S chu S 4, N 62, N 213 and A b63 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.