Distance and Orientation Dependence of Heterogeneous Electron Transfer: A Surface-Enhanced Resonance Raman Scattering Study of Cytochrome c Bound to Carboxylic Acid Terminated Alkanethiols Adsorbed on Silver Electrodes

Abstract

The distance and orientation dependence of the heterogeneous electron-transfer reaction between ferrocytochrome c (Fe2+Cc) and a silver film over nanosphere (AgFON) electrode is examined in detail using electrochemical surface-enhanced resonance Raman spectroscopy (SERRS) as a molecularly specific and structurally sensitive probe. The distance between the Fe2+ redox center and the electrode surface is controlled by varying the chain length x of an intervening carboxylic acid terminated alkanethiol, HS(CH2)xCOOH, self-assembled monolayer (SAM). The orientation of the heme in Fe2+Cc with respect to the AgFON/S(CH2)x-COOH electrode surface is controlled by its binding motif. Electrostatic binding of Fe2+Cc to AgFON/S(CH2)x-COOH yields a highly oriented redox system with the heme edge directed toward the electrode surface. The binding constants were determined to be K = 5.0 × 106 M-1 and 1.1 × 106 M-1, respectively, for the x = 5 and x = 10 SAMs. In contrast, covalent binding of Fe2+Cc yields a randomly oriented redox system with no preferred direction between the heme edge and the electrode surface. SERRS detected electrochemistry demonstrates that Fe2+Cc electrostatically bound to the x = 5 AgFON/S(CH2)xCOOH surface exhibits reversible oxidation to ferricytochrome c, whereas Fe2+Cc electrostatically bound to the x = 10 surface exhibits irreversible oxidation. In comparison, Fe2+Cc covalently bound to the x = 5 and x = 10 surfaces both exhibit oxidation with an intermediate degree of reversibility. In addition to these primary results, the work presented here shows that AgFON/S(CH2)xCOOH surfaces (1) are biocompatible - Fe2+Cc is observed in its native state and (2) are stable to supporting electrolyte changes spanning a wide range of ionic strength and pH thus enabling, for the first time, SERRS studies of these variables in a manner not accessible with either the widely used colloid or electrochemically roughened SERS-active surfaces.