Novel p16 binding peptide development for p16‐overexpressing cancer cell detection using phage display. (10th March 2015)

• Record Type: Journal Article
• Authors: Khemthongcharoen, Numfon; Ruangpracha, Athisake; Sarapukdee, Pongsak; Rattanavarin, Santi; Jolivot, Romuald; Jarujareet, Ungkarn; Plaimas, Kitiporn; Bhattarakosol, Parvapan; Patumraj, Suthiluk; Piyawattanametha, Wibool
• Is Part Of: Journal of peptide science. Volume 21:Number 4(2015:Apr.)
• Page Start: 265
• Page End: 273
• Publication Date: 2015-03-10
• Subjects: p16 -- cervical cancer -- phage display -- peptide‐protein docking -- surface alignment; Peptides -- Periodicals; Peptides -- Periodicals; 572.65
• DOI: 10.1002/psc.2726
• Languages: English
• ISSNs: 1075-2617

Abstract

Abstract : Protein p 16INK4a (p16) is a well‐known biomarker for diagnosis of human papillomavirus (HPV) related cancers. In this work, we identify novel p16 binding peptides by using phage display selection method. A random heptamer phage display library was screened on purified recombinant p16 protein‐coated plates to elute only the bound phages from p16 surfaces. Binding affinity of the bound phages was compared with each other by enzyme‐linked immunosorbent assay (ELISA), fluorescence imaging technique, and bioinformatic computations. Binding specificity and binding selectivity of the best candidate phage‐displayed p16 binding peptide were evaluated by peptide blocking experiment in competition with p16 monoclonal antibody and fluorescence imaging technique, respectively. Five candidate phage‐displayed peptides were isolated from the phage display selection method. All candidate p16 binding phages show better binding affinity than wild‐type phage in ELISA test, but only three of them can discriminate p16‐overexpressing cancer cell, CaSki, from normal uterine fibroblast cell, HUF, with relative fluorescence intensities from 2.6 to 4.2‐fold greater than those of wild‐type phage. Bioinformatic results indicate that peptide 'Ser‐His‐Ser‐Leu‐Leu‐Ser‐Ser' binds to p16 molecule with the best binding score and does not interfere with the common protein functions of p16. Peptide blocking experiment shows that the phage‐displayed peptide 'Ser‐His‐Ser‐Leu‐Leu‐Ser‐Ser' can conceal p16 from monoclonal antibody interaction. This phage clone also selectively interacts with the p16 positive cell lines, and thus, it can be applied for p16‐overexpressing cell detection. Copyright © 2015 European Peptide Society and John Wiley & Sons, Ltd. Abstract : This work identifies novel p16 binding peptides for p16‐overexpressing cancer cell detection by using phage display selection method. Binding properties of the bound phages were evaluated by enzyme‐linked immunosorbent assay (ELISA), fluorescence imaging technique, bioinformatic computation, peptide blocking experiment, and fluorescence imaging technique. The results show that phage display peptide 'Ser‐His‐Ser‐Leu‐Leu‐Ser‐Ser' is the best binding phage. This phage clone can interact selectively to p16 positive cell lines and shows the high potential to be applied for p16‐overexpressing cell detection.