Adsorptive immobilization of cytochrome c on indium/tin oxide (ITO): electrochemical evidence for electron transfer-induced conformational changes

Abstract The adsorptive immobilization and electrochemistry of horse and yeast cytochrome c on indium/tin oxide (ITO) electrodes is reported. Near-monolayer coverage was achieved in pH 7 phosphate buffers of ionic strength equal to 10 and 50 mM, respectively, for the horse and yeast species. The layers exhibit very well-behaved voltammetry and are stable on the timescale of hours to days. Cyclic voltammetry revealed quasireversible behavior that is a product of both electron transfer (ET) kinetics and ET-induced conformational changes. A square scheme mechanism linking the redox states and the conformational states is proposed. Using a simple ET kinetic model that adequately describes the voltammetry at higher scan rates, a standard ET rate constant of 18 s −1 was determined for adsorbed horse cytochrome c . With decreasing scan rate, we observed a limiting peak separation of approximately 10 mV, an example of unusual quasireversibility (UQR) that we attribute to the effect of conformational changes. Finally we note that the intrinsic cytochrome c ET rate on ITO is some 6 orders of magnitude less than for gold.

[1]  D R Dunphy,et al.  The electroactive integrated optical waveguide:  ultrasensitive spectroelectrochemistry of submonolayer adsorbates. , 1997, Analytical chemistry.

[2]  J. Chambers,et al.  Role of nucleation in the redox reactions of TTF-TCNQ single crystal electrodes in contact with aqueous KBr solutions , 1998 .

[3]  M. Erecińska,et al.  Cytochrome c interaction with membranes. Interaction of cytochrome c with isolated membrane fragments and purified enzymes. , 1974, Archives of biochemistry and biophysics.

[4]  E. F. Bowden,et al.  Voltammetric Peak Broadening for Cytochrome c/Alkanethiolate Monolayer Structures: Dispersion of Formal Potentials , 1997 .

[5]  F. Armstrong,et al.  Voltammetric characterization of rapid and reversible binding of an exogenous thiolate ligand at a [4Fe-4S] cluster in ferredoxin III from Desulfovibrio africanus , 1993 .

[6]  P. Hildebrandt,et al.  Cytochrome c at charged interfaces. 1. Conformational and redox equilibria at the electrode/electrolyte interface probed by surface-enhanced resonance Raman spectroscopy. , 1989, Biochemistry.

[7]  P. Hildebrandt,et al.  Heterogeneous Electron Transfer of Cytochrome c on Coated Silver Electrodes. Electric Field Effects on Structure and Redox Potential , 2001 .

[8]  Isao Taniguchi,et al.  The rate of conformational change of cytochrome c during electron transfer determined by double-potential step chronoelliptometry , 1990 .

[9]  R. Forster,et al.  Protonation Effects on Superexchange across Gold/Osmium Bis(bipyridyl) Tetrazine Chloride Monolayer Interfaces , 2001 .

[10]  F. M. Hawkridge,et al.  Temperature and electrolyte effects on the electron-transfer reactions of cytochrome c , 1985 .

[11]  G. Brayer,et al.  Oxidation state-dependent conformational changes in cytochrome c. , 1992, Journal of molecular biology.

[12]  Q. Chi,et al.  Ordered Assembly and Controlled Electron Transfer of the Blue Copper Protein Azurin at Gold (111) Single-Crystal Substrates , 2001 .

[13]  F. Armstrong,et al.  Fast-scan cyclic voltammetry of protein films on pyrolytic graphite edge electrodes: characteristics of electron exchange. , 1998, Analytical chemistry.

[14]  James L. Willit,et al.  Adsorption and Redox thermodynamics of strongly adsorbed cytochrome c on Tin oxide electrodes , 1990 .

[15]  Henry N. Blount,et al.  Interfacial electrochemistry of cytochrome c at tin oxide, indium oxide, gold, and platinum electrodes , 1984 .

[16]  E. Laviron.  General expression of the linear potential sweep voltammogram in the case of diffusionless electrochemical systems , 1979 .

[17]  J. Ulstrup,et al.  Voltammetry of native and recombinant Pseudomonas aeruginosa azurin on polycrystalline Au- and single-crystal Au(111)-surfaces modified by decanethiol monolayers , 2001 .

[18]  J. M. Wallace,et al.  CONTROLLING INTERFACIAL ELECTRON-TRANSFER KINETICS OF CYTOCHROME C WITH MIXED SELF-ASSEMBLED MONOLAYERS , 1998 .

[19]  K. Niki,et al.  Electroreflectance spectroscopic study of the electron transfer rate of cytochrome c electrostatically immobilized on the ω-carboxyl alkanethiol monolayer modified gold electrode , 1995 .

[20]  V. Birss,et al.  Electrochemistry of adsorbed flavin adenine dinucleotide in acidic solutions , 1991 .

[21]  James L. Willit,et al.  Determination of unimolecular electron transfer rate constants for strongly adsorbed cytochrome c on tin oxide electrodes , 1987 .

[22]  N. Tao,et al.  High-resolution multiwavelength surface plasmon resonance spectroscopy for probing conformational and electronic changes in redox proteins. , 2000, Analytical chemistry.

[23]  G. S. Wilson,et al.  Recent developments in faradaic bioelectrochemistry , 2000 .

[24]  F. Wudl,et al.  Electrochemical reduction and oxidation of C60 films , 1991 .

[25]  Isao Taniguchi,et al.  The Direct Electron Transfer Reactions of Cytochrome c at Electrode Surfaces , 1995 .

[26]  T. Kuwana,et al.  Electrochemical and Surface Characteristics of Tin Oxide and Indium Oxide Electrodes , 1976 .

[27]  T. Cotton,et al.  An electrochemical approach to investigate gated electron transfer using a physiological model system: Cytochrome c immobilized on carboxylic acid-terminated alkanethiol self-assembled monolayers on gold electrodes , 2000 .

[28]  C. Kittel.  Introduction to solid state physics , 1954 .