Properties of the periplasmic nitrate reductases from Paracoccus pantotrophus and Escherichia coli after growth in tungsten-supplemented media
Open accessAbstract
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 microM) reduced nitrate at a rate of V(max)=3.41+/-0.16 micromol [NO(3)(-)] min(-1) mg(-1) with an affinity for nitrate of K(m)=0.24+/-0.05 mM and was heat-stable up to 50 degrees C. In contrast, the periplasmic fraction obtained from cells cultured in media supplemented with tungstate (100 microM) reduced nitrate at a much slower rate, with much lower affinity (V(max)=0.05+/-0.002 micromol [NO(3)(-)] min(-1) mg(-1) and K(m)=3.91+/-0.45 mM) and was labile during prolonged incubation at >20 degrees 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.
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Tungsten can deactivate molybdenum-containing nitrate reductases by replacing the molybdenum atom at the active site.
“tungsten can deactivate these enzymes by replacing the molybdenum atom at the active site [43]”
From Nitrate reduction to nitrite, nitric oxide and ammonia by gut bacteria under physiological conditions (2015) · cites this paper as [43]
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- Authors
- Gates, A. J., Hughes, R. O., Sharp, S. R., Millington, P. D., Nilavongse, A., Cole, J. A., Leach, E.-R., Jepson, B., Richardson, D. J., & Butler, C. S.
- Affiliations
- Newcastle University (GB)
- University of Birmingham (GB)
- University of East Anglia (GB)
- Corresponding author
- Butler, Clive S
- Venue
- FEMS Microbiology Letters
- Publisher
- Oxford University Press (OUP)
- Published
- 2003-03
- Volume
- 220
- Issue
- 2
- Pages
- 261-269
- ISSN
- 0378-1097 (ISSN-L)
- 1574-6968
- OpenAlex
- W2022642927
- Cited by
- 53
- References
- 39
- Citations by year
- 2026: 3 · 2025: 1 · 2024: 2 · 2023: 4 · 2022: 1 · 2021: 4 · 2020: 2 · 2019: 4 · 2018: 2 · 2017: 1 · 2015: 3 · 2014: 1 · 2013: 1 · 2012: 4
- FWCI
- 2.28
- Type
- journal article
- Language
- en
- Open access
- bronze
- doi.org
- Locations
- FEMS Microbiology Letters (publishedVersion)
- PubMed (publishedVersion)
- University of Birmingham Research Portal (University of Birmingham) (submittedVersion)
- UEA Digital Repository (University of East Anglia) (submittedVersion)
- Newcastle University ePrints (Newcastle Univesity)
- Funders
- University of Oxford
- Biotechnology and Biological Sciences Research Council
- Topics
- Physical Sciences › Energy › Renewable Energy, Sustainability and the Environment › Metalloenzymes and iron-sulfur proteins
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Bacterial Genetics and Biotechnology
- Physical Sciences › Environmental Science › Pollution › Wastewater Treatment and Nitrogen Removal
- Keywords
- periplasmic nitrate reductase, napA, tungstate, molybdenum, tungsten substitution, molybdoenzymes, Paracoccus pantotrophus, Escherichia coli, denitrification, enzyme kinetics, enzyme stability, tungstate inhibition, anaerobic bacteria
- MeSH terms
- Anaerobiosis, Culture Media, Escherichia coli, Molybdenum, Nitrate Reductases, Nitrates, Oxidation-Reduction, Paracoccus, Tungsten, Gene Expression Regulation, Bacterial, Tungsten Compounds
About these checks
Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.
Statistics and arithmetic are recomputed, not judged.
Confidence bands are shown only for fields calibrated on our annotated test set. None is calibrated yet, so no band is shown.
What counts as a claim ›How we measure confidence ›How we check this ›
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