Catalytic protein voltammetry

Anderson, L.J., Richardson, D.J., and Butt, J.N. (2001) Catalytic protein film voltammetry from a respiratory nitrate reductase provides evidence for complex electrochemical modulation of enzyme activity. Biochemistry, 40, 11294-11307. 

When the redox activity of an adsorbed protein is coupled to catalytic redox transformation of a molecule in solution, termed the substrate, then catalytic voltammetry is observed. Electrons may be relayed between the active site and electrode by the action of one or more ancillary redox centers, as implied by the cartoon in Fig. 1. Alternatively, direct electron exchange between the electrode and the catalytic center may occur. In either case, when the electrode potential provides sufficient driving force for catalysis, there will be a sustained flow of current due to the net transfer of electrons between the electrode and molecules in solution. Fig. 4. This situation is in contrast to the discrete peaks that arise from cyclic voltammetry of non-tumover events. Figs. 2 and 3. A negative (positive) catalytic current describes catalytic reduction (oxidation). The catalytic current magnitude quantifies the rate of electron transfer through the adsorbed enzyme and so the catalytic rate. Voltammetry of a solution of the substrate should always be performed in the absence of enzyme to confirm that the catalytic currents arise from the activity of the enzyme rather than direct catalytic transformation of the substrate by the electrode surface. 

Fe 2S], a [4Fe-4S] and a [3Fe-4S] center. The enzyme catalyzes the reversible redox conversion of succinate to fumarate. Voltammetry of the enzyme on PGE electrodes in the presence of fumarate shows a catalytic wave for the reduction of fumarate to succinate (much more current than could be accounted for by the stoichiometric reduction of the protein active sites). Typical catalytic waves have a sigmoidal shape at a rotating disk electrode, but in the case of succinate dehydrogenase the catalytic wave shows a definite peak. This window of optimal potential for electrocatalysis seems to be a consequence of having multiple redox sites within the enzyme. Similar results were obtained with DMSO reductase, which contains a Mo-bis(pterin) active site and four [4Fe 4S] centers. 

Figure 4-1. Protein film voltammetry as a technique for studying redox enzyme mechanisms. The catalytic current-potential profile provides information on the rate-defining catalytic processes occurring within the enzyme. It is important that interfacial electron transfer is facile and information is not masked by limitations due to tlie transport of substrate and product for this reason the rotating disc electrode is an important tool in these studies.

As for iyrd, the combination of voltammetry and in situ STM has thus addressed a standing issue in protein film voltammetry, viz. the fraction of immobilized protein molecules that retains full functionality on surface immobilization. Two other observations illuminate this. One is that the enzyme function is more subtle and sensitive to non-native environments than simpler electron transfer function. Retaining full CuNiR catalytic... 

Protein film voltammetry (PFV) has been demonstrated in order to evaluate ET and catalytic properties of MCO directly on the electrode surface. In this approaeh, the electrode and redox centers of the enzyme are considered to be a donor-acceptor pair. The electrode donates or accepts electrons over a continuous range of potentials, and current is used as an indicator of the movement of electrons within the enzyme [36-39]. 

Hirst J. Elucidating the mechanisms of coupled electron transfer and catalytic reactions by protein film voltammetry. Biochim Biophys Acta 2006 1757 225 239. 

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