Kinetic and thermodynamic resolution of the interactions between sulfite and the pentahaem cytochrome NrfA from Escherichia coli
| Autor: | Andrew M. Hemmings, David J. Richardson, Thomas A. Clarke, Colin W. J. Lockwood, Myles R. Cheesman, Sophie J. Marritt, Susannah R. Poock, Gemma L. Kemp, Julea N. Butt |
|---|---|
| Rok vydání: | 2010 |
| Předmět: |
Models
Molecular Cytochrome biology Thiocyanate Chemistry Nitric-oxide reductase Protein Conformation Inorganic chemistry Active site Cytochrome c Group Cell Biology Heme Nitrite reductase Biochemistry Nitric oxide chemistry.chemical_compound Kinetics Sulfite Catalytic Domain biology.protein Escherichia coli Thermodynamics Nitrite Molecular Biology Oxidation-Reduction |
| Zdroj: | The Biochemical journal. 431(1) |
| ISSN: | 1470-8728 |
| Popis: | 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. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|