Direct electron transfer (DET) reactions of recombinant tobacco peroxidase (rTOP), namely direct electroreduction of Compound I/ Compound II and heme Fe3+/2+ conversion, were studied on gold electrodes. rTOP of wild type, non-glycosylated, was produced using, an Escherichia coli expression system. At pH 5.0, the redox potential for direct electrochemical transformation of the Fe3+/2+ of the peroxidase heme was -143 mV vs. Ag vertical bar AgCl, and 0.26 +/- 0.07 pmol of the adsorbed rTOP were in DET contact with the gold electrode. The total amount of the adsorbed rTOP estimated from QCM data was 53 +/- 5 pmol/cm(2) or 1.67 pmol when referred to the surface area of the electrodes used for electrochemical measurements. Of 1.67 pmol of adsorbed rTOP, only 0.76 pmol were catalytically active. DET between Au and the enzyme was also studied in the reaction of the bioelectrocatalytic reduction of H2O2 by cyclic voltammetry and amperometric detection of H2O2 at +50 mV with rTOP-modified Au electrodes placed in a wall-jet flow-through electrochemical cell. Maximal bioelectrocatalytic current response of the rTOP-modified gold electrodes to H2O2 was observed at pH 5.0 and stemmed from its bioelectrocatalytic reduction based on DET between Au and the active site of rTOP. Kinetic analysis of the DET reactions gave 52% of the adsorbed rTOP molecules active in DET reactions (0.4 pmol of adsorbed catalytically active rTOP, correspondingly), which correlated well with the non-catalytic-voltammetry data. DET was characterised by a heterogeneous ET rate constant of 13.2 s(-1), if one takes into account the QCM data, and 19.6 s(-1), if the amount of rTOP estimated from the data on DET transformation of Fe3+/2+ couple of rTOP is considered. The sensitivity for H2O2 obtained for the rTOP-modified Au electrodes was 0.7 +/- 0.1 A M-1 cm(-2). These are the first ever-reported data on DET reactions of anionic plant peroxidases on bare gold electrodes. (c) 2005 Elsevier B.V. All rights reserved.
Bibliographical noteThe information about affiliations in this record was updated in December 2015.
The record was previously connected to the following departments: Analytical Chemistry (S/LTH) (011001004)
Subject classification (UKÄ)
- Analytical Chemistry
- Impedance spectroscopy
- Quartz crystal microbalance
- Heterogeneous direct electron transfer
- Gold electrodes
- Recombinant tobacco peroxidase