ISBN: 978-83-60043-10-3 - eurobic9

Eurobic9, 2-6 September, 2008, Wrocław, Poland
P165. Electron Paramagnetic Resonance as a Tool for Investigating how
Redox Active Enzymes Attach to the Surfaces of Electrodes
Maxie M. Roessler, a Jeffrey Harmer, a Fraser A. Armstrong a
a Inorganic Chemistry Laboratory, Department of Chemistry, University of Oxford, South Parks Road, Oxford
OX1 3QR, United Kingdom
Electron paramagnetic resonance (EPR) is being applied to redox-enzymes bound to conductive surfaces to
probe the interaction between them. Such surfaces can be thought of as replacing the physiological redox partner
of the enzyme and are the basis of the technique called protein film voltammetry (PFV) for studying enzymes
[1]. PFV has been used extensively to gain fundamental information about the active sites of enzymes, to control
their catalytic action, to measure absolute turn-over frequencies in the absence of substrate-diffusion limitations,
and to quantify the rate of electron transfer within the redox-active protein [2,3].
Pyrolytic graphite ‘edge’ has been shown to be a versatile electrode surface for binding enzymes. After abrasive
pre-treatment it is very rough and rich in C-O functionalities. When measured with a small molecule (N2), the
actual surface area of a typical electrode is in fact 10 4 times greater than its visible geometric surface area [4].
However, to date little is known about the way an enzyme interacts with such a surface, which is important for
further development of the technique. For instance, it may be that only a fraction of the total number of adsorbed
enzymes on the surface are electrochemically active, that is, only a fraction of the enzyme molecules are in the
‘correct’ orientation for electrons from the electrode to tunnel through the protein to the active site.
Enzyme-modified micro- and nano-particles of the electrode material, in particular graphite, have been produced
and explored with EPR. First investigations have been carried out on laccase from Trametes versicolor, which
possesses four copper centres, two of which are paramagnetic and can be probed directly. A schematic
illustration is given in the figure below.
Cartoon of an enzyme attached to an electrode as studied with PFV and its extension to ‘electrode particles’
modified with enzyme that is a suitable arrangement for EPR experiments.
Acknowledgements: The authors acknowledge the EPSRC (EP/D048559/1) for support of this research and St.
John’s College Oxford for a travel grant.
References:
[1] Leger, C.; Elliott, S. J.; Hoke, K. R.; Jeuken, L. J. C.; Jones, A. K.; Armstrong, F. A. Biochemistry 2003, 42,
8653-8662.
[2] Vincent, K. A.; Armstrong, F. A. Inorg. Chem. 2005, 44, 798-809.
[3] Vincent, K. A.; Parkin, A.; Armstrong, F. A. Chem. Rev. (Washington, DC, U. S.) 2007, 107, 4366-4413.
[4] Blanford, C. F.; Armstrong, F. A. J. Solid State Electrochem. 2006, 10, 826-832.
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