The Direct Electrochemistry of Redox-active Proteins

The term direct electrochemistry of proteins means the possibility to detect the direct exchange of electrons between the active site(s) of a protein and a (metallic or inert material) electrode without the help of redox mediators, which might favour an indirect interaction between the electrode and the protein (see the discussion on Electrocatalysis in Chapter 2, Section 1.4.4). This aspect of electrochemistry is not yet as widely explored as it deserves, but the relevant results are now analysed in a rather comprehensive fashion.1  

Other biological redox processes are for instance important in the bacterial degradation of chemical products found in the soil (such as the oxygenases illustrated in Chapter 9, Section 1.1). 

As electrochemistry is particularly suitable for studying electron transfer events, its application to biological redox processes appears reasonable. Unfortunately, the difficulties encountered and the few results obtained until the beginning of the 1980s meant that it became firmly believed that it would not be possible to use this technique to study the direct electron transfers activated by proteins. The basis of this scepticism were the following  

In biological systems the movement of an electron from a donor to an acceptor site is a quite common and apparently simple event. In reality, however, electron transfers over distances greater than 10 A are frequently necessary, which means that the electron transfer can be a slow process. In fact, according to the Marcus theory, the rate constant for electron transfer between two redox sites, ket (s-1), is given by 5 

A = structural reorganization energy (eV) necessary such that the donor and acceptor sites have geometrical arrangements able to favour the electron transfer  

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Since the establishment of spectroelectrochemistry very little effort has been devoted to the direct electrochemistry of redox proteins. Although many thermodynamic and kinetic parameters can be determined by UV-VIS spectroelectrochemistry, the electrochemical reaction mechanisms for redox proteins are not well understood. New techniques md new theoretical treatments are needed to address this issue. Moreover, most attention has been placed on relatively simple electron transfer proteins to date no one has reported the direct electrochemistry of a more complex system (e.g., a redox enzyme system) which unequivocally undergoes electron transfer to (or from) its active site. Considerable experimental work is needed to develop more fully spectroelectrochemical methods for biological systems. 

The first reports on direct electrochemistry of a redox active protein were published in 1977 by Hill [49] and Kuwana [50], They independently reported that cytochrome c (cyt c) exhibited virtually reversible electrochemistry on gold and tin doped indium oxide (ITO) electrodes as revealed by cyclic voltammetry, respectively. Unlike using specific promoters to realize direct electrochemistry of protein in the earlier studies, recently a novel approach that only employed specific modifications of the electrode surface without promoters was developed. From then on, achieving reversible, direct electron transfer between redox proteins and electrodes without using any mediators and promoters had made great accomplishments. 

The opportunity of obtaining direct electrochemistry of cytochrome c and other metalloproteins at various electrode materials such as modified gold and pyrolytic graphite has led to numerous reports of heterogeneous electron transfer rates and mechanisms between the protein and the electrode. In all the reports, Nicholson s method (37) was employed to calculate rate constants, which were typically within the range of 10" -10 cm sec with scan rates varying between 1 and 500 mV sec This method is based on a macroscopic model of the electrode surface that assumes that mass transport of redox-active species to and from the electrode occurs via linear diffusion to a planar disk electrode and that the entire surface is uniformly electroactive, i.e., the heterogeneous electron transfer reaction can take place at any area. 

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