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Gates, Andrew J. 
ORCID: https://orcid.org/0000-0002-4594-5038, Hughes, Richard O., Sharp, Sarah R., Millington, Paul D., Nilavongse, Arjaree, Cole, Jeffrey A., Leach, Emily-Rose, Jepson, Brian J. N., Richardson, David J. ORCID: https://orcid.org/0000-0002-6847-1832 and Butler, Clive S.
(2003)
Properties of the periplasmic nitrate reductases from Paracoccus pantotrophus and Escherichia coli after growth in tungsten-supplemented media.
FEMS Microbiology Letters, 220 (2).
pp. 261-269.
ISSN 1574-6968
Abstract
Paracoccus pantotrophus grown anaerobically under denitrifying conditions expressed similar levels of the periplasmic nitrate reductase (NAP) when cultured in molybdate- or tungstate-containing media. A native PAGE gel stained for nitrate reductase activity revealed that only NapA from molybdate-grown cells displayed readily detectable nitrate reductase activity. Further kinetic analysis showed that the periplasmic fraction from cells grown on molybdate (3 µM) reduced nitrate at a rate of Vmax=3.41±0.16 µmol [NO3-] min-1 mg-1 with an affinity for nitrate of Km=0.24±0.05 mM and was heat-stable up to 50°C. In contrast, the periplasmic fraction obtained from cells cultured in media supplemented with tungstate (100 µM) reduced nitrate at a much slower rate, with much lower affinity (Vmax=0.05±0.002 µmol [NO3-] min-1 mg-1 and Km=3.91±0.45 mM) and was labile during prolonged incubation at >20°C. Nitrate-dependent growth of Escherichia coli strains expressing only nitrate reductase A was inhibited by sub-mM concentrations of tungstate in the medium. In contrast, a strain expressing only NAP was only partially inhibited by 10 mM tungstate. However, none of the above experimental approaches revealed evidence that tungsten could replace molybdenum at the active site of E. coli NapA. The combined data show that tungsten can function at the active site of some, but not all, molybdoenzymes from mesophilic bacteria.
| Item Type: | Article |
|---|---|
| Faculty \ School: | Faculty of Science > School of Biological Sciences |
| UEA Research Groups: | Faculty of Science > Research Groups > Molecular Microbiology Faculty of Science > Research Centres > Centre for Molecular and Structural Biochemistry Faculty of Science > Research Groups > Organisms and the Environment |
| Related URLs: | |
| Depositing User: | Users 2731 not found. |
| Date Deposited: | 15 Jun 2011 11:07 |
| Last Modified: | 16 May 2023 08:30 |
| URI: | https://ueaeprints.uea.ac.uk/id/eprint/32589 |
| DOI: | 10.1016/S0378-1097(03)00122-8 |
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