Kinetic and thermodynamic resolution of the interactions between sulfite and the pentahaem cytochrome NrfA from Escherichia coli

Kemp, Gemma L., Clarke, Thomas A. ORCID: https://orcid.org/0000-0002-6234-1914, Marritt, Sophie J., Lockwood, Colin, Poock, Susannah R., Hemmings, Andrew M. ORCID: https://orcid.org/0000-0003-3053-3134, Richardson, David J. ORCID: https://orcid.org/0000-0002-6847-1832, Cheesman, Myles R. and Butt, Julea N. ORCID: https://orcid.org/0000-0002-9624-5226 (2010) Kinetic and thermodynamic resolution of the interactions between sulfite and the pentahaem cytochrome NrfA from Escherichia coli. Biochemical Journal, 431 (1). pp. 73-80. ISSN 0264-6021

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Abstract

NrfA is a pentahaem cytochrome present in a wide-range of ?-, d- and e-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.

Item Type:	Article
Faculty \ School:	Faculty of Science > School of Biological Sciences Faculty of Science > School of Chemistry
UEA Research Groups:	Faculty of Science > Research Groups > Molecular Microbiology Faculty of Science > Research Groups > Plant Sciences Faculty of Science > Research Groups > Biophysical Chemistry (former - to 2017) Faculty of Science > Research Groups > Organisms and the Environment Faculty of Science > Research Groups > Chemistry of Light and Energy Faculty of Science > Research Groups > Chemistry of Life Processes Faculty of Science > Research Centres > Centre for Molecular and Structural Biochemistry Faculty of Science > Research Groups > Energy Materials Laboratory
Depositing User:	Vishal Gautam
Date Deposited:	12 Sep 2011 10:52
Last Modified:	15 May 2023 23:56
URI:	https://ueaeprints.uea.ac.uk/id/eprint/20547
DOI:	10.1042/BJ20100866

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