Affiliation:
1. Divisions ofMicrobiology1 and
2. Laboratory Medicine and Pathobiology,2 North York, Ontario M4N 3M5, Canada
3. Dermatology,3 Sunnybrook and Women's College Health Sciences Centre, and
4. University of Toronto Departments of Medicine4 and
Abstract
ABSTRACT
A fibronectin (Fn)-binding adhesin of
Staphylococcus aureus
contains three tandem 37- or 38-amino-acid motifs (D1, D2, and D3), which function to bind Fn. Plasma from patients with
S. aureus
infections contain antibodies that preferentially recognize ligand induced binding sites in the D motifs and do not inhibit Fn binding (F. Casolini, L. Visai, D. Joh, P. G. Conaldi, A. Toniolo, M. Höök, and P. Speziale, Infect. Immun. 66:5433–5442, 1998). To eliminate the influence of Fn binding on antibody development, we used synthetic peptide immunogens D1
21–34
and D3
20–33
, which each contain a conserved pattern of amino acids that is essential for Fn binding but which cannot bind Fn without N- or C-terminal extensions. The D3
20–33
immunogen promoted the production of polyclonal antibodies that were 10-fold more effective as inhibitors of Fn-binding to the D3 motif than antibodies obtained by immunizing with an extended peptide D3
16–36
, which exhibits functional Fn binding. The D3
20–33
immunogen also facilitated the production of a monoclonal antibody, 9C3, which was highly specific for the epitope SVDFEED, and abolished Fn binding by the D3 motif. When mixed with polyclonal anti-D1
21–34
immunoglobulin G, 70% inhibition of Fn binding to the three tandem D motifs was achieved compared to no more than 30% inhibition with either antibody preparation alone. Therefore, by immunizing with short synthetic peptides that are unable to bind Fn, we have effectively stimulated the production of antibodies specific for epitopes comprised of amino acids that are essential for Fn binding. Although these epitopes occur within a conserved pattern of amino acids that is required for Fn binding, the antibodies recognized specific linear epitope sequences and not a conserved structure common to all repeated motifs.
Publisher
American Society for Microbiology
Subject
Infectious Diseases,Immunology,Microbiology,Parasitology
Cited by
23 articles.
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