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Peptide Scanning-assisted Identification of a Monoclonal Antibody-recognized Linear B-cell Epitope
Published on: March 24, 2017
Determination of the Binding Epitope for Anti-trichosanthin Monoclonal Antibody T(8)C(12)
Hui-Dong Yuan1, Mo-Yi Li, Xiao-Xia Shao
1Shanghai Institute of Biochemistry, the Chinese Academy of Sciences, Shanghai 200031, China.
Western blot result showed that T(8)C(12), an anti-trichosanthin (TCS) monoclonal antibody, could bind to a CNBr-cleaved TCS fragment with MW of 8 kD. The epitope was located in 1-72 of the N terminal of TCS as shown by amino acid analysis. A random 6-aa peptide library cloned in pIII of phage M13 was screened by T(8)C(12). After two cycles screening, 15 positive clones were randomly selected and six kinds of 6-aa sequences were determined, which showed to be highly homologous with a Ser/Thr-(X)-(X)-Arg motif, the X denoting hydrophobic amino acids. The motif was found to be similar to the first 3-5 amino acid sequence of the TCS N-terminal. The synthesized 1-8 peptide of TCS showed competitive binding to T(8)C(12) with native TCS. It suggested that 3-5 amino acid residues of TCS N-terminal was the core of epitope recognized by T(8)C(12).
Western blot result showed that T(8)C(12), an anti-trichosanthin (TCS) monoclonal antibody, could bind to a CNBr-cleaved TCS fragment with MW of 8 kD. The epitope was located in 1-72 of the N terminal of TCS as shown by amino acid analysis. A random 6-aa peptide library cloned in pIII of phage M13 was screened by T(8)C(12). After two cycles screening, 15 positive clones were randomly selected and six kinds of 6-aa sequences were determined, which showed to be highly homologous with a Ser/Thr-(X)-(X)-Arg motif, the X denoting hydrophobic amino acids. The motif was found to be similar to the first 3-5 amino acid sequence of the TCS N-terminal. The synthesized 1-8 peptide of TCS showed competitive binding to T(8)C(12) with native TCS. It suggested that 3-5 amino acid residues of TCS N-terminal was the core of epitope recognized by T(8)C(12).
