Identification and Characterization of Novel Binding Epitope of Tetanus toxoid by Phage Display Peptide Library

Order of Publishing in Issue: 
7
Volume :9
Issue :1
February, 2015
Page No: 
49-58
Authors: 
Neelakantam Biradhar,[1,2] Sridevi V Nimmagadda,[1] Shukra M Aavula,[1] Sugumar Parthasarathy,[1] Samuel Sula1 and Kapil Maithal[1*]
Address: 
[1]Research and Development Center, Indian Immunologicals Limited, Rakshapuram, Gachibowli, Hyderabad-500 032, Telangana, India.
Address: 
[2]Department of Biotechnology, Acharya Nagarjuna University, Guntur-522510, Andhra Pradesh, India.

Phage display peptide libraries are widely used in various protein-protein interaction studies to determine the immunological binding of proteins and epitope mapping of different targets. In the present study, the peptide library is used to identify the Fab binding epitope site of Tetanus Toxoid (TT). The peptides were screened against Fab by biopanning, using a random peptide 12- mer phage display library. Clones were selected based on the binding activity by phage ELISA and one of the peptide sequence (DTMSYTPNIHLL), which showed highest binding activity towards the Fab was cloned into pGEX-4T1 vector for expression and purified using glutathione agarose affinity chromatography. The binding activity of the purified peptide was demonstrated by ELISA and Immunoblot analysis. Further, analysis of the peptide sequence revealed homology of four amino acids (YTPN) to heavy chain sequence of TT. F13 Fab significantly binds to the YTPN epitope of the TT antigen as indicated by ELISA and may have potential in diagnostics.

Keywords: 
Tetanus toxoid, Fab, Phage display, Peptide, ELISA.
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