Generating short peptidic ligands for silver nanowires from phagedisplay random librariesPriscilla Chan, ? Tiffany Phan, Michael C. Kao, Cheryl Dolan and Jeffrey B.-H. Tok *Department of Chemistry and Materials Science, Biosecurity and Nanosciences Laboratory, Lawrence Livermore National Laboratory,University of California, Livermore, CA 94551, USAReceived 5 June 2006; revised 25 July 2006; accepted 1 August 2006Available online 24 August 2006Abstract—We report the generation of peptide ligands for silver nanowires using a linear 12-mer peptide phage display randomlibrary technique. Phage clones that specifically bind the silver nanowires are sequenced after three rounds of biopanning, andobtained DNA sequences suggest that there are a few conserved amino acid residues which may be critical for binding. A selectedbinding peptide, together with two mutant peptide sequences, were subsequently synthesized on Tentagel resins to examine theimportance of both the identities and positions of the conserved amino acid residues.? 2006 Elsevier Ltd. All rights reserved.Phage display random library is an established and use-ful tool to rapidly generate binding peptide ligandsagainst a wide variety of targets in biomedical, physical,and chemical research. 1–7 A particularly exciting field isthe application of the phage library against nano- andmicroparticles. In specific, there have been a series ofelegant work by Belcher and co-workers regarding thegeneration of peptide sequences for the self-assemblyof semi-conducting nanoparticles. 8–11 In addition, Stoneand co-workers have reported the usage of phage dis-play library against acid-etched AgNP for in vitro bio-synthesis of AgNP through Ag-binding peptides. 12,13As our long-term interest lies in the usage of biomole-cules for patterning and/or self-assembly of metallicnanowires, we seek to address the ‘universality’ of pep-tide sequences against Ag structures with different mor-phologies, for example, spherical particles versus wires.AgNW are obtained from Nanoplex Technologies Inc.(Mountain View, CA), in which their dimensions are250 nm in diameter and 6 lm in length. The AgNWare supplied in H 2 O, with a concentration of ?10 9 nano-wires per milliliter. The linear 12-mer phage display pep-tides library kit is obtained from New England Biolabs(Medford, MA). The library is generated from filamen-tous Escherichia coli phage M13, with the random pep-tide portion being fused to N-terminal of the minor coatproteins pIII. The complexity of the library is reportedto contain ?1.28 · 10 9 independent sequences. Thelibrary was maintained and amplified with E. coli strainER2738, which was provided along with the phagedisplay library kit.As for the biopanning process, ?10 3 AgNW are beingplaced in a microfuge tube, washed twice, and resus-pended in 100 lL of binding buffer (100 lL TBS–0.1%Tween 20). Subsequently, 10–20 lL of the library stocksolution (?2 · 10 12 phage) in 1 mL of binding buffer wasadded, and incubated, with mixing, for 1 h at room tem-perature. The microfuge tube was then centrifuged at5000 rpm for 1 min to pellet the AgNW, and the un-bound phage was carefully removed along with thesupernatant. The AgNW-phage pellet was then washedfive times by repeated resuspension and centrifugationof AgNW in 500 lL of washing buffer (TBS–0.1%Tween 20). Finally, the bound phage was eluted by add-ing 100 lL of 0.2 M glycine–HCl, pH 2.2, containing0.1% BSA. After 10 min of incubation at room temper-ature, the mixture was neutralized with 15 lL of 1 MTris–HCl, pH 9.1. The neutralized mixture containing0960-894X/$ - see front matter ? 2006 Elsevier Ltd. All rights reserved.doi:10.1016/j.bmcl.2006.08.013Abbreviations: AgNW, silver nanowires; AgNP, silver nanoparticles;X-Gal, 5-bromo-4-chloro-3-indolyl-b- D -galactoside; IPTG, isopropyl-b- D -thiogalactoside.Keywords: Phage display random library; Nanowires; Aminoacid residues, Tentagel resins; In vitro biosynthesis. .* Corresponding author. Tel.: +1 925 423 1549; fax: +1 925 4223570; e-mail: tok2@llnl.gov?Present address: Department of Pharmacology, Columbia University,New York, NY 10032, USA.Bioorganic & Medicinal Chemistry Letters 16 (2006) 5261–5264
