Epitope Mapping Using Ribosome Display in a Reconstituted Cell-Free Protein Synthesis System

Authors: Osada, Eriko; Shimizu, Yoshihiro; Akbar, Bintang K.; Kanamori, Takashi; Ueda, Takuya

Source: The Journal of Biochemistry, Volume 145, Number 5, 27 May 2009 , pp. 693-700(8)

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Abstract:

Ribosome display is a powerful technology for selecting ligand-binding peptides or proteins. We demonstrate here that the ribosome display using the reconstituted cell-free protein synthesis system can be applied for the epitope mapping of monoclonal antibodies (mAbs). Using this technology, we selected peptides that specifically bind to three mAbs from random peptide library. When selection was performed against the anti-FLAG M2 antibody, selected peptides contained previously characterized consensus epitope, indicating that the methodology can be applied for the epitope mapping. When the selection was carried out against two anti-β-Catenin (anti-β-Cat) mAbs, selected peptides had a homology for the partial peptide sequences of β-Cat. Western blot analysis showed that these putative epitopes had affinity for the corresponding mAbs and β-Cat mutants that lack these regions did not bind to the antibodies, indicating we correctly mapped the epitope for these mAbs. The study shown here provides a way for the quick identification of the epitope of mAbs.

Keywords: antibody; cell-free protein synthesis system; epitope mapping; in vitro selection; ribosome display

Document Type: Regular paper

DOI: http://dx.doi.org/10.1093/jb/mvp027

Publication date: 2009-05-27