Protein Film, and Diffusion–Controlled Voltammetry of Hemeproteins on a Single Surface

Abstract

Protein film voltammetry has made possible electrochemical studies of complex protein reactions, such as redox-induced conformational dynamics1 and enzyme kinetics.2 However, without diffusion-controlled experiments as a reference, one cannot be sure of the extent to which the protein–electrode interaction affects conformational dynamics or enzyme kinetics. Diffusion-controlled voltammetric experiments can provide an alternate view into these processes. Previously, semiconductor electrodes have been successfully employed for diffusion-controlled experiments.3 Herein, we report novel application of electrochemically deposited cadmium oxide (CdO) as a working electrode for both diffusion-controlled voltammetry as well as protein-film voltammetry (PFV). The hydrophilic CdO surface allows transient protein interactions for diffusion-controlled measurements, as well as a suitable surface for drop casting protein films. Polycrystalline CdO electrodes were electrodeposited onto a fluorine-doped tin oxide (FTO) working electrode held at –0.7 V vs. SCE for 15 min in aqueous solutions of CdCl2, then annealed at 450 °C for 2 h. CdO surfaces were characterized by means of x-ray photoelectron spectroscopy (XPS), x-ray diffraction (XRD), and scanning electron microscopy (SEM). XPS and XRD confirm formation of polycrystalline CdO. Electron micrographs confirm complete coverage of the FTO by CdO. To generate protein films, 10 mg/mL solutions of either cytochrome c or hemoglobin were drop cast onto CdO electrodes and left for 2 h. Cyclic voltammograms recorded for both hemoglobin and cytochrome c display nearly reversible electron transfer, with redox potentials similar to those previously reported.4 1 A. El Kasmi, M. C. Leopold, R. Galligan, R. T. Robertson, S. S. Saavedra, K. El Kacemi, E. F. Bowden, Electrochem. Commun. 2002, 4, 177–181. 2 H. Liu, G. Zou, J. Biochem. Biophys. Methods. 2006, 68, 87–99. 3 G. Valincius, V. Reipa, M. P. Mayhew, V. L. Walker, Electrochem. Commun. 2002, 4, 314–317. 4 G. Battistuzzi, M. Bosari, M. Sola, F. Francia, Biochemistry, 1997, 36, 16247–16258.