pH and Substrate Effect on Adsorption of Peptides Containing Z and E Dehydrophenylalanine. Surface-Enhanced Raman Spectroscopy Studies on Ag Nanocolloids and Electrodes Kamilla Malek,* ,† Agata Kró likowska, ‡ and Jolanta Bukowska ‡ † Faculty of Chemistry, Jagiellonian University, Ingardena 3, 30-060 Krakow, Poland ‡ Faculty of Chemistry, University of Warsaw, Pasteura 1, 02-093 Warsaw, Poland * S Supporting Information ABSTRACT: The silver substrates and pH dependent surface-enhanced Raman scattering (SERS) spectra of unsaturated derivatives of di- and tripeptides (dehydropeptides) are investigated. Experimental spectra were interpreted with the help of DFT calculations and normal-mode analysis. We choose as objects of our studies modified but natural peptides containing Z or E dehydrophenylalanine ( (Z)/(E) ΔPhe) residue to study an effect of the type of the isomer on the interaction between the peptide and silver surfaces in the form of nanocolloidal particles and an electrochemically roughened electrode. We also observed that the SERS profile is sensitive to both the type of the studied SERS active substrate and pH, especially for the adsorption on the silver colloid. In general, all dehydropeptides interact with both SERS substrates upon deprotonation of the C-end of the molecule. The participation of the other fragments of the adsorbates such as the N-terminal amino group and the dehydroresidue is also manifested in the SERS spectra. Their orientation with respect to the silver surfaces is discussed in detail. 1. INTRODUCTION Normal Raman and surface-enhanced Raman spectroscopy is a powerful tool in the characterization of biologically active species such as proteins, nucleic acids, drugs, etc. 1 Both techniques are sensitive to the even slight changes in the molecular structure, as well as the chemical environment. SERS refers to the extraordinarily large enhancement of Raman signals that are obtained from roughened surfaces or aggregated small nanoparticles of certain metals, usually silver, gold, and copper. The most used SERS substrates are metal electrodes and colloidal nanoparticles because of their low costs as well as a simple manipulation. Obviously, roughness of a SERS substrate affects the adsorption mechanism of a molecule by changing strength of the metal-adsorbate interactions and/or an adsorption site. Despite this, Coulombic stabilization between the metal surface and the adsorbate is required to observe surface-enhanced Raman scattering; thus when the electrode surface or colloidal particles and the analyte have charges of the same sign, the adsorption process can be strongly hindered. Consequently, an orientation of the adsorbate on the metal as well as detection level of an analyte can be completely different, when various metal substrates are used in SERS experiment. 1 Hence, the investigation of the substrate influence, its interfacial properties, and molecular specificity is crucial for a better understanding of the SERS enhancement for a given molecule. The Raman profile of amino acids, peptides, and proteins, especially those adsorbed on the metal surface investigated in SERS experiment, has been widely studied by several groups. 2-7 Because of the large Raman cross-section of the aromatic moieties, SERS spectra of aromatic peptides or proteins are dominated by bands assigned to vibrations of the phenyl ring. The latter is accompanied by the bands of the deprotonated C-terminal end of a biomolecule and rarely by modes of the amino groups or the peptide backbone. In turn, introducing α,β-dehydrophenylalanine (ΔPhe) residue into the backbone of a peptide sequence affects both chemical reactivity and conformation, because this residue contains a double C-C bond between the α and β carbon atoms. The presence of this bond decreases the conformational flexibility of proteins. Additionally, the ΔPhe moiety exists in the two isomeric forms, Z and E, which can influence the biological activity of the peptide. 8 This class of nonproteino- genic amino acids can be found in many biological peptides with antiviral, antitumor, anti-inflammatory, or immunosup- pressive activities. 9,10 Dehydroamino acids are also used in de novo synthesis of protein mimics for structure-function relationship studies. 11,12 In addition, potentiometric and electronic absorption studies on coordination ability of dehydropeptides toward Ni(II) and Cu(II) ions have shown that the presence of the double C α C β bond makes the dehydropeptides more efficient ligands than the parent peptides. 13,14 These dehydropeptides form various coordina- Received: January 20, 2014 Revised: March 13, 2014 Article pubs.acs.org/JPCB © XXXX American Chemical Society A dx.doi.org/10.1021/jp500650p | J. Phys. Chem. B XXXX, XXX, XXX-XXX