Identification of Brucella cell wall antigenic determinants by immunoscreening of a random peptide library expressed at the surface of filamentous phage

Summary

Antibodies against the 89 kDa Brucella abortus outer membrane protein (OMP) are detected in 68% of B. abortus infected cows. A monoclonal antibody, specifically directed against Brucella OMP89, has been used to screen a phage-displayed peptide library. We describe here results obtained from affinity selection of phages with mAb A7617E3C3 in three rounds of biopanning using a cysteine-constrained peptide library expressed on the surface of filamentous bacteriophages pVIII major coat protein. Deduced amino acid sequences of the peptide region of clones positively detected by colony immunoblotting experiments indicate the presence of a consensus sequence. Peptides corresponding to the most frequently represented sequence have been synthesized. Monoclonal antibody binding to selected phages is inhibited by corresponding cyclic peptides, but not by linear peptides. This confirms the specificity of the peptide sequence for its paratope but also the importance of a certain conformation for binding.