Format

Send to

Choose Destination
J Pept Sci. 2015 Apr;21(4):265-73. doi: 10.1002/psc.2726. Epub 2015 Mar 10.

Novel p16 binding peptide development for p16-overexpressing cancer cell detection using phage display.

Author information

1
NECTEC, National Science and Technology Development Agency (NSTDA), Pathumthani, 12120, Thailand; Advanced Imaging Research Center, Faculty of Medicine, Chulalongkorn University, Bangkok, 10330, Thailand.

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.

KEYWORDS:

cervical cancer; p16; peptide-protein docking; phage display; surface alignment

PMID:
25754556
DOI:
10.1002/psc.2726
[Indexed for MEDLINE]

Supplemental Content

Loading ...
Support Center