Periplasmic nitrate reduction in Wolinella succinogenes: cytoplasmic NapF facilitates NapA maturation and requires the menaquinol dehydrogenase NapH for membrane attachment
Abstract
Various nitrate-reducing bacteria produce proteins of the periplasmic nitrate reductase (Nap) system to catalyse electron transport from the membraneous quinol pool to the periplasmic nitrate reductase NapA. The composition of the corresponding nap gene clusters varies but, in addition to napA, genes encoding at least one membrane-bound quinol dehydrogenase module (NapC and/or NapGH) are regularly present. Moreover, some nap loci predict accessory proteins such as the iron–sulfur protein NapF, whose function is poorly understood. Here, the role of NapF in nitrate respiration of the Epsilonproteobacterium Wolinella succinogenes was examined. Immunoblot analysis showed that NapF is located in the membrane fraction in nitrate-grown wild-type cells whereas it was found to be a soluble cytoplasmic protein in a napH deletion mutant. This finding indicates the formation of a membrane-bound NapGHF complex that is likely to catalyse NapH-dependent menaquinol oxidation and electron transport to the iron–sulfur adaptor proteins NapG and NapF, which are located on the periplasmic and cytoplasmic side of the membrane, respectively. The cysteine residues of a CX 3 CP motif and of the C-terminal tetra-cysteine cluster of NapH were found to be required for interaction with NapF. A napF deletion mutant accumulated the catalytically inactive cytoplasmic NapA precursor, suggesting that electron flow or direct interaction between NapF and NapA facilitated NapA assembly and/or export. On the other hand, NapA maturation and activity was not impaired in the absence of NapH, demonstrating that soluble NapF is functional. Each of the four tetra-cysteine motifs of NapF was modified but only one motif was found to be essential for efficient NapA maturation. It is concluded that the NapGHF complex plays a multifunctional role in menaquinol oxidation, electron transfer to periplasmic NapA and maturation of the cytoplasmic NapA precursor.
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Nap has the ability to function in nitrate respiration.
“Similar to Nar, Nap has the ability to function in nitrate respiration.”
From Nitrate and periplasmic nitrate reductases (2014) · cites this paper as [103]
Mutation of W. succinogenes NapF leads to accumulation of the inactive precursor NapA in the cytoplasm.
“Moreover mutation of the W. succinogenes NapF led to the accumulation of the inactive precursor NapA in the cytoplasm.103”
From Nitrate and periplasmic nitrate reductases (2014) · cites this paper as [103]
The gene napL encodes a 35 kDa periplasmic chaperone protein with no known metal binding motifs.
“The gene napL encodes a 35 kDa periplasmic chaperone protein,103,105 with no known metal binding motifs.103”
From Nitrate and periplasmic nitrate reductases (2014) · cites this paper as [103]
One specific motif of NapF is responsible for transferring the [4Fe-4S] cluster into NapA.
“Mutation of each motif demonstrated that one specific motif is responsible for transferring the [4Fe–4S] cluster into NapA.103”
From Nitrate and periplasmic nitrate reductases (2014) · cites this paper as [103]
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- Authors
- Kern, M., & Simon, J.
- Affiliations
- Goethe University Frankfurt (DE)
- ORCID
- Simon, Jörg
- Venue
- Microbiology
- Publisher
- Microbiology Society
- Published
- 2009-08-01
- Volume
- 155
- Issue
- 8
- Pages
- 2784-2794
- ISSN
- 1350-0872 (ISSN-L)
- 1465-2080
- OpenAlex
- W1970719212
- Cited by
- 33
- References
- 33
- Citations by year
- 2025: 1 · 2023: 1 · 2022: 1 · 2021: 1 · 2019: 2 · 2018: 5 · 2017: 2 · 2016: 3 · 2015: 6 · 2014: 2 · 2013: 2 · 2012: 1
- FWCI
- 1.36
- Type
- journal article
- Language
- en
- Open access
- closed
- Locations
- Microbiology (publishedVersion)
- PubMed (publishedVersion)
- Funders
- Deutsche Forschungsgemeinschaft
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Bacterial Genetics and Biotechnology
- Physical Sciences › Energy › Renewable Energy, Sustainability and the Environment › Metalloenzymes and iron-sulfur proteins
- Physical Sciences › Environmental Science › Environmental Engineering › Microbial Fuel Cells and Bioremediation
- Keywords
- Wolinella succinogenes, periplasmic nitrate reductase, Napf, nitrate respiration, electron transport, iron-sulfur proteins, protein assembly, Epsilonproteobacteria
- MeSH terms
- Bacterial Outer Membrane Proteins, Cytoplasm, Multigene Family, Nitrate Reductases, Nitrates, Oxidation-Reduction, Wolinella, Periplasm, Amino Acid Motifs, Multiprotein Complexes
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