Epitope mapping using ribosome display in a reconstituted cell-free protein synthesis system.
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-beta-Catenin (anti-beta-Cat) mAbs, selected peptides had a homology for the partial peptide sequences of beta-Cat. Western blot analysis showed that these putative epitopes had affinity for the corresponding mAbs and beta-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.
Osada
Eriko
E
The Department of Medical Genome Sciences, Graduate School of Frontier Sciences, the University of Tokyo, Kashiwa, Chiba, Japan.
Shimizu
Yoshihiro
Y
Akbar
Bintang K
BK
Kanamori
Takashi
T
Ueda
Takuya
T
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000276:immunology; D015153:Blotting, Western; D002474:Cell-Free System; D018604:Epitope Mapping; D008969:Molecular Sequence Data; D050505:Mutant Proteins / Q000378:metabolism; D019151:Peptide Library; D010455:Peptides / Q000737:chemistry; D014176:Protein Biosynthesis; D012270:Ribosomes / Q000378:metabolism*; D020539:Sequence Analysis, Protein; D051176:beta Catenin / Q000276:immunology
D000911:Antibodies, Monoclonal; D050505:Mutant Proteins; D019151:Peptide Library; D010455:Peptides; D051176:beta Catenin