PMID:20629638
| Citation |
Kemp, GL, Clarke, TA, Marritt, SJ, Lockwood, C, Poock, SR, Hemmings, AM, Richardson, DJ, Cheesman, MR and Butt, JN (2010) Kinetic and thermodynamic resolution of the interactions between sulfite and the pentahaem cytochrome NrfA from Escherichia coli. Biochem. J. 431:73-80 |
|---|---|
| Abstract |
NrfA is a pentahaem cytochrome present in a wide-range of γ-, δ- and ε-proteobacteria. Its nitrite and nitric oxide reductase activities have been studied extensively and contribute to respiratory nitrite ammonification and nitric oxide detoxification respectively. Sulfite is a third substrate for NrfA that may be encountered in the micro-oxic environments where nrfA is expressed. Consequently, we have performed quantitative kinetic and thermodynamic studies of the interactions between sulfite and Escherichia coli NrfA to provide a biochemical framework from which to consider their possible cellular consequences. A combination of voltammetric, spectroscopic and crystallographic analyses define dissociation constants for sulfite binding to NrfA in oxidized (~54 μM), semi-reduced (~145 μM) and reduced (~180 μM) states that are comparable with each other, and the Km (~70 μM) for sulfite reduction at pH 7. Under comparable conditions Km values of ~22 and ~300 μM describe nitrite and nitric oxide reduction respectively, whereas the affinities of nitrate and thiocyanate for NrfA fall more than 50-fold on enzyme reduction. These results are discussed in terms of the nature of sulfite co-ordination within the active site of NrfA and their implications for the cellular activity of NrfA. |
| Links |
PubMed Online version:10.1042/BJ20100866 |
| Keywords |
Catalytic Domain; Cytochrome c Group/chemistry; Cytochrome c Group/metabolism; Escherichia coli/enzymology; Escherichia coli/metabolism; Heme/chemistry; Heme/metabolism; Kinetics; Models, Molecular; Oxidation-Reduction; Protein Conformation; Thermodynamics |
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