Electrochemistry of Redox Enzymes

Direct, unmediated electrochemistry of redox enzymes has interested many researchers in several aspects. Understanding of the thermodynamics, kinetics, stoichiometry, and interfacial properties of redox enzymes is obviously important. The most attractive aspect, however, is the use of enzyme electrodes as novel electrochemical biosensors and their applications to bioreactors and biofuel cells. Although the observation of direct electrochemistry of small redox proteins has become almost commonplace as the consequence of extensive research over the past decade, the corresponding study with larger redox enzymes has proved more elusive. The difficulty lies mainly in that the redox centers are located sufficiently far from the outermost 

There appear to be two classes of redox enzymes intrinsic and extrinsic (5). With the former, the catalytic reaction between an enzyme and its substrates takes place within a highly localized assembly of redox-active sites. There need be no electron transfer pathways from these sites to the surface of the enzyme, where, it is presumed, it would interact with an electrode. For such intrinsic redox enzymes, electrode reactions may require (1) that the sites of the catalytic reaction be close to the protein surface, (2) that the enzyme can deform without loss of activity, (3) that the electrode surface projects into the enzyme. 

The first accounts that seemed to give direct enzyme electrochemistry were the reports concerning a soluble blue Cu oxidase, laccase, which catalyzed the rapid four-electron reduction of dioxygen to water. An efficient electrocatalysis of O2 reduction by adsorbed fungal laccase on pyrolytic graphite, glassy carbon, and C02-treated carbon black electrodes was first described by Tarasevich and co-workers (48). Several control experiments were carried out to verify direct electron transfer from the electrode to the Cu sites of the enzyme. 

Another example of enzyme electrochemistry is given by cytochrome c peroxidase (C( P). This monomer molecule contains a 5-type heme and catalyzes the two-electron reduction of H2O2 to water using cytochrome c (II) as the electron donor. In the process, the Fe(III) center of the enzyme reacts rapidly with H2O2 to yield a two-electron oxidized species, compound I, which is reduced to the Fe(III) form via another species, compound II. 

Direct electron transfer between CCP and an electrode was first reported (45) for the nonphysiological one-electron reduction and reoxidation of ferric CCP at fluorine-doped tin oxide. Overpotentials of around 0.5 V were required to drive this electrode reaction in either direction at measurable rates. A more successful approach to direct electroreduction of compound I, described by Armstrong and Lannon (46), employed edge-plane graphite electrodes in the presence of 

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C. The Microscopic Model in Protein Electrochemistry Electrochemistry of Protein-Protein Complexes Electrochemistry of Redox Enzymes... 

Leger, C. and Bertrand, P. (2008) Direct electrochemistry of redox enzymes as a tool for mechanistic studies. Chemical Reviews, 108 (7), 2379-2438. 

Rusling JF, Wang B, Yun S (2008) Electrochemistry of redox enzymes. In Bartlett PN (ed) Bioelectrochemistry, fundamentals, experimental techniques and applications. Wiley, Hoboken... 

Nowadays, studies of direct electrochemistry of redox proteins at the electrodesolution interface have held more and more scientists interest. Those studies are a convenient and informative means for understanding the kinetics and thermodynamics of biological redox processes. And they may provide a model for the study of the mechanism of electron transfer between enzymes in biological systems, and establish a foundation for fabricating new kinds of biosensors or enzymatic bioreactors. 

The determination of H202 is very important in many different fields, such as in clinical, food, pharmaceutical, and environmental analyses [202], Many techniques such as spectrophotometry, chemiluminesence, fluorimetry, acoustic emission, and electrochemistry methods have been employed to determine H202. Electrochemical methods are often used because of their advantages. Among these electrochemical methods, the construction of the mediator-free enzyme-based biosensors based on the direct electrochemistry of redox proteins has been reported over the past decade [203— 204], The enzyme-based biosensors, which use cyt c as biocatalyzer to catalyze H202, were widely studied. 

The use of redox enzymes in organic synthesis, while having a large potential for broad application in the selective formation of high-value compounds, has been limited by the necessity of cofactor regeneration or enzyme reactivation. Electrochemistry offers an attractive and, in principle, simple way to solve this problem because the mass-free electrons are used as regenerating agents. No... 

The effect of mutations or chemical modification is mostly measured in terms of changes in peroxidase activity. However, the electron transfer rate depends on the redox potential of the enzyme, so that a more integral characterization should include the electrochemistry of the enzyme. Thus, the modulation of redox potential could also serve for the design of more efficient biocatalysts. 

The direct electrochemistry of redox proteins has developed significantly in the past few years. Conditions now exist that permit the electrochemistry of all the proteins to be expressed at a range of electrodes, and important information about thermodynamic and kinetic properties of these proteins can be obtained. More recently, direct electron transfer between redox enzymes and electrodes has been achieved due to the more careful control of electrode surfaces. The need for biocompatible surfaces in bioelectrochemistry has stimulated the development of electrode surface engineering techniques, and protein electrochemistry has been reported at conducting polymer electrodes 82) and in membranes 83, 84). Furthermore, combination of direct protein electrochemistry with spectroscopic methods may offer 85) a novel way of investigating structure-function relationships in electron transport proteins. 

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. 

Moreover, within the recent developments in Bioelectrochemistry, relevant attention has been devoted to the electrochemistry of redox proteins and redox enzymes (H.A.O. Hill), as welt as of synthetic models of copper proteins (J.O. Cabral). 

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