Publication Type Journal Article
Title Analysis of the activation mechanism of Pseudomonas stutzeri cytochrome c peroxidase through an electron transfer chain
Authors Patricia M. Paes de Sousa D. Rodrigues C. G. Timoteo M. L. Simoes Goncalves G. W. Pettigrew I. Moura Margarida Correia dos Santos
Groups
Journal JOURNAL OF BIOLOGICAL INORGANIC CHEMISTRY
Year 2011
Month August
Volume 16
Number 6
Pages 881-888
Abstract The activation mechanism of Pseudomonas stutzeri cytochrome c peroxidase (CCP) was probed through the mediated electrochemical catalysis by its physiological electron donor, P. stutzeri cytochrome c-551. A comparative study was carried out, by performing assays with the enzyme in the resting oxidized state as well as in the mixed-valence activated form, using cyclic voltammetry and a pyrolytic graphite membrane electrode. In the presence of both the enzyme and hydrogen peroxide, the peak-like signal of cytochrome c-551 is converted into a sigmoidal wave form characteristic of an E(r)C(i) catalytic mechanism. An intermolecular electron transfer rate constant of (4 +/- 1) x 10(5) M(-1) s(-1) was estimated for both forms of the enzyme, as well as a similar Michaelis-Menten constant. These results show that neither the intermolecular electron transfer nor the catalytic activity is kinetically controlled by the activation mechanism of CCP in the case of the P. stutzeri enzyme. Direct enzyme catalysis using protein film voltammetry was unsuccessful for the analysis of the activation mechanism, since P. stutzeri CCP undergoes an undesirable interaction with the pyrolytic graphite surface. This interaction, previously reported for the Paracoccus pantotrophus CCP, induces the formation of a non-native conformation state of the electron-transferring haem, which has a redox potential 200 mV lower than that of the native state and maintains peroxidatic activity.
DOI http://dx.doi.org/10.1007/s00775-011-0785-8
ISBN
Publisher
Book Title
ISSN 0949-8257
EISSN
Conference Name
Bibtex ID ISI:000292887500007
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