title: Isolation, kinetic analysis, and structural characterization of an antibody targeting the Bacillus anthracis major spore surface protein BclA.
abstract: One method of laboratory- or field-based testing for anthrax is detection of Bacillus anthracis spores by high-affinity, high specificity binding reagents. From a pool of monoclonal antibodies, we selected one such candidate (A4D11) with high affinity for tBclA, a truncated version of the B. anthracis exosporium protein BclA. Kinetic analysis utilising both standard and kinetic titration on a Biacore biosensor indicated antibody affinities in the 300 pM range for recombinant tBclA, and the A4D11 antibody was also re-formatted into scFv configuration with no loss of affinity. However, assays against B. anthracis and related Bacilli species showed limited binding of intact spores as well as significant cross-reactivity between species. These results were rationalized by determination of the three-dimensional crystallographic structure of the scFv-tBclA complex. A4D11 binds the side of the tBclA trimer, contacting a face of the antigen normally packed against adjacent trimers within the exosporium structure; this inter-spore interface is highly conserved between Bacilli species. Our results indicate the difficulty of generating a high-affinity antibody to differentiate between the highly conserved spore structures of closely related species, but suggest the possibility of future structure-based antibody design for this difficult target.
authors.affiliation: CSIRO Division of Materials Science and Engineering, Parkville, Victoria, 3052, Australia. Stewart.Nuttall@csiro.au
authors.forename: Stewart D
authors.initials: SD
authors.lastname: Nuttall
authors.forename: Michelle L
authors.initials: ML
authors.lastname: Wilkins
authors.forename: Victor A
authors.initials: VA
authors.lastname: Streltsov
authors.forename: Luisa
authors.initials: L
authors.lastname: Pontes-Braz
authors.forename: Olan
authors.initials: O
authors.lastname: Dolezal
authors.forename: Hung
authors.initials: H
authors.lastname: Tran
authors.forename: Chun-Qiang
authors.initials: CQ
authors.lastname: Liu
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000907:Antibodies, Bacterial / Q000737:chemistry* / Q000276:immunology / Q000378:metabolism; D000911:Antibodies, Monoclonal / Q000737:chemistry* / Q000276:immunology / Q000378:metabolism; D001408:Bacillus anthracis / Q000737:chemistry / Q000276:immunology*; D001426:Bacterial Proteins / Q000276:immunology* / Q000378:metabolism; D015374:Biosensing Techniques; D018360:Crystallography, X-Ray; D006825:Hybridomas; D008562:Membrane Glycoproteins / Q000276:immunology* / Q000378:metabolism; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D057127:Single-Chain Antibodies / Q000737:chemistry* / Q000276:immunology / Q000378:metabolism
chemical_list: D000907:Antibodies, Bacterial; D000911:Antibodies, Monoclonal; D001426:Bacterial Proteins; C470593:BclA protein, Bacillus anthracis; D008562:Membrane Glycoproteins; D057127:Single-Chain Antibodies