2010/07/15 by Gemma L. Kemp, Gemma L. Kemp, Thomas A. Clarke +15
Biochemistry, Genetics and Molecular Biology · Energy · #Metabolomics and Mass Spectrometry Studies #Metalloenzymes and iron-sulfur proteins #Sulfur Compounds in Biology
paper · pdf · doi:10.1042/bj20100866
openalex publication_date 2010/07/15 · openalex created_date 2016/06/24 · openalex updated_date 2026/07/28
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.