title: Structural basis of anthrax edema factor neutralization by a neutralizing antibody.
abstract: Fine epitope mapping of EF13D, a highly potent neutralizing monoclonal antibody specific for the anthrax edema factor (EF), was accomplished through random mutagenesis and yeast surface display. A yeast-displayed library of single point mutants of an EF domain III (DIII), comprising amino acids 624-800, was constructed by random mutagenesis and screened for reduced binding to EF13D. With this method, residues Leu 667, Ser 668, Arg 671, and Arg 672 were identified as key residues important for EF13D binding. They form a contiguous patch on a solvent-exposed surface at one end of the four-helix bundle of DIII. Computational protein-protein docking experiments between anEF13D model and a crystal structure of EF indicate that the EF13D heavy chain complementarity-determining region 3 (HCDR3) is deeply buried within a hydrophobic cleft between two helices of DIII and interacts directly with residues Leu 667, Ser 668, Arg 671 and Arg 672, providing an explanation for the high binding affinity. In addition, they show that the HCDR3 binding site overlaps with the binding site of the N-terminal lobe of calmodulin (CaM), an EF enzymatic activator, consistent with a previous finding showing direct competition with CaM that results in neutralization of EF. Identifying the neutralization epitope of EF13D on EF improves our understanding of the neutralization mechanism and has implications for vaccine development.
authors.affiliation: Laboratory of Infectious Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, United States.
authors.forename: Michelle
authors.initials: M
authors.lastname: Makiya
authors.forename: Michael
authors.initials: M
authors.lastname: Dolan
authors.forename: Liane
authors.initials: L
authors.lastname: Agulto
authors.forename: Robert
authors.initials: R
authors.lastname: Purcell
authors.forename: Zhaochun
authors.initials: Z
authors.lastname: Chen
mesh_terms: D000595:Amino Acid Sequence; D022122:Anthrax Vaccines / Q000276:immunology; D000911:Antibodies, Monoclonal / Q000737:chemistry* / Q000235:genetics / Q000276:immunology*; D057134:Antibodies, Neutralizing / Q000737:chemistry* / Q000235:genetics / Q000276:immunology*; D000915:Antibody Affinity; D000942:Antigens, Bacterial / Q000276:immunology; D001427:Bacterial Toxins / Q000037:antagonists & inhibitors* / Q000276:immunology; D018604:Epitope Mapping; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009500:Neutralization Tests; D011487:Protein Conformation
chemical_list: D022122:Anthrax Vaccines; D000911:Antibodies, Monoclonal; D057134:Antibodies, Neutralizing; D000942:Antigens, Bacterial; D001427:Bacterial Toxins; C030325:anthrax toxin