Spectroelectrochemical characterization of a pentaheme cytochrome in solution and as electrocatalytically active films on nanocrystalline metal-oxide electrodes.
The pentaheme containing cytochrome, NrfA, from Escherichia coli catalyzes the six-electron reduction of nitrite and the five-electron reduction of nitric oxide. Crystallographic and spectroscopic studies have provided a structural framework for these mechanisms. The active site includes a high-spin heme, and four low-spin, bis-his coordinated hemes are positioned to facilitate intra- and intermolecular electron exchange. However, despite the use of protein film voltammetry to provide kinetic descriptions of NrfA catalysis at graphite and gold electrodes, the thermodynamic descriptions of heme redox activity remain incomplete. Here we rectify this situation with the observation of nonturnover signals from NrfA adsorbed on mesoporous SnO2 electrodes. Simultaneous cyclic voltammetry and electronic absorption spectroscopy define reduction potentials for the high- and low-spin hemes. These reduction potentials are shown to be similar to those exhibited by the enzyme in solution and defined by electrodic reduction monitored by magnetic circular dichroism. Thus, NrfA is shown to undergo minimal perturbation of its electronic and thermodynamic properties on adsorption giving confidence to correlations of properties deduced from various methods and in approaches that may well facilitate studies of other oxidoreductases where catalytic protein film voltammetry is well-defined but nonturnover signals elusive.
Centre for Metalloprotein Spectroscopy and Biology, School of Chemical Sciences and Pharmacy, School of Biological Sciences, University of East Anglia, Norwich Research Park, Norwich, NR4 7TJ, UK.
Sophie J
SJ
Marritt
Gemma L
GL
Kemp
Li
L
Xiaoe
James R
JR
Durrant
Myles R
MR
Cheesman
Julea N
JN
Butt
D002384:Catalysis; D003574:Cytochrome c Group / Q000737:chemistry*; D004563:Electrochemistry; D004566:Electrodes; D053758:Nanoparticles / Q000737:chemistry*; D010084:Oxidation-Reduction; D012996:Solutions; D013057:Spectrum Analysis; D013816:Thermodynamics; D017971:Tin Compounds / Q000737:chemistry*
D003574:Cytochrome c Group; D012996:Solutions; D017971:Tin Compounds; C022302:cytochrome C-552; C045358:stannic oxide