Resolution of two native monomeric 90kDa nitrate reductase active proteins from Shewanella gelidimarina and the sequence of two napA genes
Abstract
The reduction of nitrate to nitrite in the bacterial periplasm occurs in the 90kDa NapA subunit of the periplasmic nitrate reductase (NAP) system. Most Shewanella genomes contain two nap operons: napEDABC and napDAGHB, which is an unusual feature of this genus. Two native, monomeric, 90kDa nitrate reductase active proteins were resolved by hydrophobic interaction chromatography from aerobic cultures of Shewanella gelidimarina replete with reduced nitrogen compounds. The 90kDa protein obtained in higher yield was characterized as NapA by electronic absorption and electron paramagnetic resonance spectroscopies and was identified by LC/MS/MS and MALDI-TOF/TOF MS as NapA from the napEDABC-type operon. The other 90kDa protein, which was unstable and produced in low yields, was posited as NapA from the napDAGHB-type operon. Two napA genes have been sequenced from the napEDABC-type and napDAGHB-type operons of S. gelidimarina. Native NAP from S. putrefaciens was resolved as one NapA monomer and one NapAB heterodimer. Two amino acid substitutions in NapA correlated with the isolation of NAP as a NapA monomer or a NapAB heterodimer. The resolution of native, redox-active NapA isoforms in Shewanella provides new insight into the respiratory versatility of this genus, which has implications in bioremediation and the assembly of microbial fuel cells.
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The different dissociation constants could explain why E. coli NapA was isolated as a monomer while the R. sphaeroides homologue was isolated as a heterodimer.
“This could explain why the E. coli NapA was isolated as a monomer, while the R. sphaeroides homologue was isolated as a heterodimer.254”
From Nitrate and periplasmic nitrate reductases (2014) · cites this paper as [254]
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- Authors
- Simpson, P. J. L., McKinzie, A. A., & Codd, R.
- Affiliations
- The University of Sydney (AU)
- Corresponding author
- Codd, Rachel
- Venue
- Biochemical and Biophysical Research Communications
- Publisher
- Elsevier BV
- Published
- 2010-07
- Volume
- 398
- Issue
- 1
- Pages
- 13-18
- ISSN
- 0006-291X (ISSN-L)
- OpenAlex
- W2063501065
- Cited by
- 20
- References
- 25
- Citations by year
- 2026: 3 · 2022: 2 · 2021: 1 · 2018: 2 · 2017: 2 · 2015: 1 · 2014: 2 · 2013: 3 · 2012: 2
- FWCI
- 2.16
- Type
- journal article
- Language
- en
- Open access
- closed
- Locations
- Biochemical and Biophysical Research Communications (publishedVersion)
- PubMed (publishedVersion)
- Topics
- Physical Sciences › Environmental Science › Environmental Engineering › Microbial Fuel Cells and Bioremediation
- 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
- Keywords
- periplasmic nitrate reductase, napA, nitrate reduction, protein purification, amino acid substitutions, Shewanella putrefaciens, bioremediation, microbial fuel cells
- MeSH terms
- Amino Acid Sequence, Genes, Bacterial, Molecular Sequence Data, Operon, Mass Spectrometry, Sequence Alignment, Amino Acid Substitution, Shewanella, Nitrate Reductase
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