Phage display selection of a Pb(II) specific peptide and its application as a biorecognition unit for colorimetric detection of Pb(II) ions.

Abstract

Phage display technology employs a library of engineered filamentous M13 viruses infecting only bacteria. In this study, the genuine phage display selection technique was applied to identify a Pb(II) specific peptide. After three rounds of positive selection against Pb(II) coated agarose-based beads and the consecutive negative screenings against interfering metal ions (Al(III), Co(II), Fe(III), Ni(II), and Zn(II)), a final phage library with enhanced Pb(II) binding was obtained. Enzyme Linked Immunosorbent Assay (ELISA) analyses confirmed the selective Pb(II) binding of the enriched viruses. Twenty monoclonal phage plaques were randomly selected, single stranded DNAs (ssDNAs) were isolated and sequenced. Sequencing results revealed four different peptide sequences. Pb9 peptide (KASPYIT) showing the most specific Pb(II) binding was selected for detection studies. Pb9 was synthetically synthesized with additional three cysteine (3xCys) units at C-terminal. Twenty nanometers AuNPs were functionalized with Pb9-3xCys peptides through Au-thiol (Au-S) interaction. A colorimetric Pb(II) detection system was validated using the engineered peptide-AuNP complex at a calculated LOD of around 11nM (3σ/k, n=6) for the case study. The detection system was Pb(II) selective over various metal ions (Ag(II), Al(III), Au(III), Cd(II), Co(II), Cr (III), Cu(II), Fe(III), Hg(II), Mg(II), Mn(II), Ni(II), and Zn(II)). Such metal ion specific peptides can be further studied to develop simple, user friendly and cost-effective tools to design alternative detection and bioremediation systems for a circular economy.