title: Electrochemistry of metalloproteins: protein film electrochemistry for the study of E. coli [NiFe]-hydrogenase-1.
abstract: Protein film electrochemistry is a technique which allows the direct control of redox-active enzymes, providing particularly detailed information on their catalytic properties. The enzyme is deposited onto a working electrode tip, and through control of the applied potential the enzyme activity is monitored as electrical current, allowing for direct study of inherent activity as electrons are transferred to and from the enzyme redox center(s). No mediators are used. Because the only enzyme present in the experiment is bound at the electrode surface, gaseous and liquid phase inhibitors can be introduced and removed whilst the enzyme remains in situ. Potential control means that kinetics and thermodynamics are explored simultaneously; the kinetics of a reaction can be studied as a function of potential. Steady-state catalytic rates are observed directly as current (for a given potential) and non-steady-state rates (such as interconversions between different forms of the enzyme) are observed from the change in current with time. The more active the enzyme, the higher the current and the better the signal-to-noise. In this chapter we outline the practical aspects of PFE for studying electroactive enzymes, using the Escherichia coli [NiFe]-hydrogenase 1 (Hyd-1) as an example.
authors.affiliation: Inorganic Chemistry Laboratory, University of Oxford, South Parks Road, Oxford, OX1 3QR, UK.
authors.forename: Rhiannon M
authors.initials: RM
authors.lastname: Evans
authors.forename: Fraser A
authors.initials: FA
authors.lastname: Armstrong
mesh_terms: D055664:Electrochemical Techniques; D004563:Electrochemistry / Q000379:methods*; D004566:Electrodes; D004926:Escherichia coli / Q000201:enzymology*; D006108:Graphite / Q000737:chemistry; D006864:Hydrogenase / Q000737:chemistry / Q000378:metabolism*; D007700:Kinetics; D008667:Metalloproteins / Q000378:metabolism*; D010084:Oxidation-Reduction
chemical_list: D008667:Metalloproteins; D006108:Graphite; C043296:nickel-iron hydrogenase; D006864:Hydrogenase