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Direct electrochemistry and biolelectrocatalysis of H2O2 reduction of recombinant tobacco peroxidase on graphite. Effect of peroxidase single-point mutation on Ca2+-modulated catalytic activity

Castillo, John LU ; Ferapontova, Elena LU ; Hushpulian, D; Tasca, Federico LU ; Tishkov, V; Chubar, T; Gazaryan, I and Gorton, Lo LU (2006) In Journal of Electroanalytical Chemistry and Interfacial Electrochemistry 588(1). p.112-121
Abstract
Direct electron transfer (DET) reactions and bio(electro)catalytic reduction of H2O2 catalysed by native and recombinant forms of tobacco peroxidase (nTOP and rTOP) were studied in homogeneous-phase catalysis and when TOPs were adsorbed on graphite electrodes. Non-glycosylated wild type and Glu141 -> Phe mutant forms of rTOP were produced using an Escherichia coli expression system. Mutation was introduced to explore the mechanisms for modulation of the catalytic activity of TOP by Ca2+ ions. At the pH optimum of 5.0, direct electrochemical Fe3+/2+ transformation of the peroxidase heme was characterised by potentials of -208 mV (nTOP) and -239 mV vs. Ag vertical bar AgCl (rTOP), and 0.9 +/- 0.1 and 1.1 +/- 0.4 pmoles of adsorbed nTOP... (More)
Direct electron transfer (DET) reactions and bio(electro)catalytic reduction of H2O2 catalysed by native and recombinant forms of tobacco peroxidase (nTOP and rTOP) were studied in homogeneous-phase catalysis and when TOPs were adsorbed on graphite electrodes. Non-glycosylated wild type and Glu141 -> Phe mutant forms of rTOP were produced using an Escherichia coli expression system. Mutation was introduced to explore the mechanisms for modulation of the catalytic activity of TOP by Ca2+ ions. At the pH optimum of 5.0, direct electrochemical Fe3+/2+ transformation of the peroxidase heme was characterised by potentials of -208 mV (nTOP) and -239 mV vs. Ag vertical bar AgCl (rTOP), and 0.9 +/- 0.1 and 1.1 +/- 0.4 pmoles of adsorbed nTOP and rTOP, correspondingly, were in DET contact with graphite. Kinetic analysis of amperometric (at +50 mV) data on H2O2 reduction at TOP-modified electrodes, placed in a wall-jet flow-through electrochemical cell, yielded 82% (nTOP) and 88% (rTOP) of adsorbed TOP molecules active in the DET reaction. The efficiency of DET (and bioelectrocatalysis) increased 3.5-fold when changing from glycosylated nTOP to rTOP. The Glu141 Phe mutation in the heme-binding pocket of rTOP enabled to achieve a Ca2+-tolerance of TOP in the reaction with H2O2, which is characteristic of other plant peroxidases, and to a large extent in heterogeneous DET and reaction with a second substrate catechol. The results promote further applications of TOP for biosensor- and solid-phase biocatalysts development. (c) 2005 Elsevier B.V. All rights reserved. (Less)
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author
organization
publishing date
type
Contribution to journal
publication status
published
subject
keywords
heterogeneous direct electron transfer, graphite electrodes, refolding, single-point mutation, peroxidase, recombinant tobacco, enzymatic catalysis, bioelectrocatalysis
in
Journal of Electroanalytical Chemistry and Interfacial Electrochemistry
volume
588
issue
1
pages
112 - 121
publisher
Elsevier
external identifiers
  • wos:000235264000013
  • scopus:31344465504
ISSN
0022-0728
DOI
10.1016/j.jelechem.2005.12.010
language
English
LU publication?
yes
id
33288013-9dab-460e-9f91-bba252bbd546 (old id 417783)
date added to LUP
2007-10-01 17:22:04
date last changed
2019-09-04 02:59:45
@article{33288013-9dab-460e-9f91-bba252bbd546,
  abstract     = {Direct electron transfer (DET) reactions and bio(electro)catalytic reduction of H2O2 catalysed by native and recombinant forms of tobacco peroxidase (nTOP and rTOP) were studied in homogeneous-phase catalysis and when TOPs were adsorbed on graphite electrodes. Non-glycosylated wild type and Glu141 -> Phe mutant forms of rTOP were produced using an Escherichia coli expression system. Mutation was introduced to explore the mechanisms for modulation of the catalytic activity of TOP by Ca2+ ions. At the pH optimum of 5.0, direct electrochemical Fe3+/2+ transformation of the peroxidase heme was characterised by potentials of -208 mV (nTOP) and -239 mV vs. Ag vertical bar AgCl (rTOP), and 0.9 +/- 0.1 and 1.1 +/- 0.4 pmoles of adsorbed nTOP and rTOP, correspondingly, were in DET contact with graphite. Kinetic analysis of amperometric (at +50 mV) data on H2O2 reduction at TOP-modified electrodes, placed in a wall-jet flow-through electrochemical cell, yielded 82% (nTOP) and 88% (rTOP) of adsorbed TOP molecules active in the DET reaction. The efficiency of DET (and bioelectrocatalysis) increased 3.5-fold when changing from glycosylated nTOP to rTOP. The Glu141 Phe mutation in the heme-binding pocket of rTOP enabled to achieve a Ca2+-tolerance of TOP in the reaction with H2O2, which is characteristic of other plant peroxidases, and to a large extent in heterogeneous DET and reaction with a second substrate catechol. The results promote further applications of TOP for biosensor- and solid-phase biocatalysts development. (c) 2005 Elsevier B.V. All rights reserved.},
  author       = {Castillo, John and Ferapontova, Elena and Hushpulian, D and Tasca, Federico and Tishkov, V and Chubar, T and Gazaryan, I and Gorton, Lo},
  issn         = {0022-0728},
  keyword      = {heterogeneous direct electron transfer,graphite electrodes,refolding,single-point mutation,peroxidase,recombinant tobacco,enzymatic catalysis,bioelectrocatalysis},
  language     = {eng},
  number       = {1},
  pages        = {112--121},
  publisher    = {Elsevier},
  series       = {Journal of Electroanalytical Chemistry and Interfacial Electrochemistry},
  title        = {Direct electrochemistry and biolelectrocatalysis of H2O2 reduction of recombinant tobacco peroxidase on graphite. Effect of peroxidase single-point mutation on Ca2+-modulated catalytic activity},
  url          = {http://dx.doi.org/10.1016/j.jelechem.2005.12.010},
  volume       = {588},
  year         = {2006},
}