Identification of linear epitopes in Bacillus anthracis protective antigen bound by neutralizing antibodies

J Biol Chem. 2009 Sep 11;284(37):25077-86. doi: 10.1074/jbc.M109.022061. Epub 2009 Jul 18.

Abstract

Protective antigen (PA), the binding subunit of anthrax toxin, is the major component in the current anthrax vaccine, but the fine antigenic structure of PA is not well defined. To identify linear neutralizing epitopes of PA, 145 overlapping peptides covering the entire sequence of the protein were synthesized. Six monoclonal antibodies (mAbs) and antisera from mice specific for PA were tested for their reactivity to the peptides by enzyme-linked immunosorbent assays. Three major linear immunodominant B-cell epitopes were mapped to residues Leu(156) to Ser(170), Val(196) to Ile(210), and Ser(312) to Asn(326) of the PA protein. Two mAbs with toxin-neutralizing activity recognized two different epitopes in close proximity to the furin cleavage site in domain 1. The three-dimensional complex structure of PA and its neutralizing mAbs 7.5G and 19D9 were modeled using the molecular docking method providing models for the interacting epitope and paratope residues. For both mAbs, LeTx neutralization was associated with interference with furin cleavage, but they differed in effectiveness depending on whether they bound on the N- or C-terminal aspect of the cleaved products. The two peptides containing these epitopes that include amino acids Leu(156)-Ser(170) and Val(196)-Ile(210) were immunogenic and elicited neutralizing antibody responses to PA. These results identify the first linear neutralizing epitopes of PA and show that peptides containing epitope sequences can elicit neutralizing antibody responses, a finding that could be exploited for vaccine design.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Animals
  • Antibodies / chemistry
  • Antigens, Bacterial / chemistry*
  • B-Lymphocytes / metabolism
  • Bacillus anthracis / metabolism*
  • Bacterial Toxins / chemistry*
  • Enzyme-Linked Immunosorbent Assay
  • Epitope Mapping
  • Epitopes / chemistry
  • Female
  • Hybridomas / metabolism
  • Macrophages / metabolism
  • Mice
  • Mice, Inbred BALB C
  • Molecular Sequence Data
  • Neutralization Tests
  • Protein Structure, Tertiary

Substances

  • Antibodies
  • Antigens, Bacterial
  • Bacterial Toxins
  • Epitopes
  • anthrax toxin

Associated data

  • GENBANK/FJ784743
  • GENBANK/FJ784744
  • GENBANK/FJ784745
  • GENBANK/FJ784746
  • GENBANK/FJ784747
  • GENBANK/FJ784748
  • GENBANK/FJ784749
  • GENBANK/FJ784750