Photophysical Probes of a Protein/Semiconductor Electrode Interface

Abstract

Immobilization of redox molecules on solid electrodes offers a way of investigating the electron transfer of a donorlacceptor interface without the presence of diffusion processes.‘ Such interactions also have important implications for analytical and clinical biochemistry. Direct electrochemistry of proteins and enzymes is therefore an active research field.2 One example of direct protein electrochemistry relevant to this work is cytochrome c (Cyt c ) adsorbed on a tin oxide (SnO2) surface.lb This protein is rich in lysine amino acid residues and thus carries positive charges at neutral pH. It has been found that the native protein, Fe-Cyt c, adsorbs strongly on SnO2, presumably through the electrostatic interaction with the deprotonated hydroxyl groups on SnOz surface (Scheme 1A). The electron transfer rate constant between the two at zero free energy has been measured by cyclic voltammetry and is found to be relatively slow (3-8 s-l). In terms of modern theoretical understanding of electron transfer? the parameters (electronic coupling, V, and reorganization energy, A) which govern this rate remain uncharacterized. We report here a novel approach to obtain such parameters by combining thermal and photochemical interfacial electron transfer measurements within the context of a novel theoretical formalism. We used time-resolved fluorescence spectroscopy to measure the interfacial electron transfer rate constant from zinc-substituted cyt c (Zn-Cyt c) to SnOz and estimated electronic coupling between the protein and semiconductor and the reaction reorganization energy. The lowest photoexcited state of Zn-Cyt c has a much higher energy level4 than the conduction band edge5 of SnO2 (Scheme IB). Electron injection to SnO2 (route a in Scheme 1B) is therefore a thermodynamically favorable process that competes with fluorescent decay emission (route b). Zn-Cyt c was prepared6 and adsorbedlbs7 onto a fluorinedoped SnO2 substrate according to the published procedures. The adsorption was confirmed by the UV-vis spectrum of ZnCyt c and cyclic voltammogram of the native Fe-Cyt c. Integration of either the anodic or the cathodic wave in the voltammogram yielded a surface coverage of 510 pmol/cm2, which is substantially less than the calculated full monolayer coverage of about 20 pmol/cm2, by assuming the diameter of