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

Eurobic9, 2-6 September, 2008, Wrocław, Poland
P134. Electrocatalytic Aldehyde Oxidation and Acid Reduction by
Hyperthermophilic Tungsten-containing Oxidoreductase on Ferredoxin-
Modified Gold
M. Nahid Hasan a , S. de Vries a , W.R. Hagen a , H.A. Heering b
a.
Department of Biotechnology, Delft University of Technology, Julianalaan 67, 2628 BC Delft, The
Netherlands.
b.
Leiden Institute of Chemistry, Leiden University, Einsteinweg 55, 2333 CC Leiden, The Netherlands.
The hyperthermophilic aldehyde:ferredoxin oxidoreductase (AOR) and glyceraldehyde-3-phosphate
(GAP):ferredoxin oxidoreductase (GAPOR) from Pyrococcus furiosus form complexes with their native redox
partner ferredoxin (Fd), chemisorbed on a gold electrode. With GAPOR, a well-developed non-turnover
voltammetric response is observed at room temperature and pH 7.4, assigned to the [4Fe 4S] clusters of the
enzyme and Fd, at 368mV and 383 mV, respectively. At 60ºC a catalytic oxidation wave is observed upon
addition of the substrate GAP. With AOR, a broad, reversible non-turnover voltammetric response is observed at
room temperature due to overlapping potentials of the Fd and AOR [4Fe-4S] clusters, in addition to
tungstopterin species. The AOR / Fd complex formation on the electrode is corroborated by ellipsometric
detection of Fd-labeled gold on immobilized AOR. At 80ºC and in the presence of the substrate crotonaldehyde,
three distinct catalytic oxidation responses are observed. Remarkably, these responses are peak-shaped due to a
rapid switch-off at high overpotentials, and are each accompanied by a peak due to the catalytic reduction of the
produced acid. The averages of the oxidation and reduction peaks are found at 0.38 V, 0.27 V and 0.10 V. The
bi-directional AOR activity and potential-dependent switching are confirmed by colorimetric aldehyde analysis
and dye mediated optical activity assays. A minimal mechanism is proposed, involving reversible productinduced
switching between an aldehyde oxidizing form and an acid reducing form of the enzyme. The present
work opens the way for the application of Fd electrodes in achieving controlled reduction of carboxylic acids on
a preparative scale.
References:
[1] Hasan MN, Kwakernaak C, Sloof WG, Hagen WR, Heering HA (2006) J Biol Inorg Chem 11:651-662
[2] Koehler BP, Mukund S, Conover RC, Dhawan I K, Roy R, Adams MWW, Johnson MK (1996) J Am Chem
Soc 118:12391-12405
[3] Mukund S, Adams MWW (1991) J Biol Chem 266:14208-14216
[4] Hagedoorn P-L, Freije JR, Hagen WR (1999) FEBS Lett 462:66-70
[5] van den Ban ECD, Willemen HM, Wassink H, Laane C, Haaker H (1999) Enzyme Microb Tech 25:251-257
[6] Bernhardt PV (2006) Aust J Chem 59:233-256
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