Unraveling an FNR based regulatory circuit in Paracoccus denitrificans using a proteomics-based approach

Bouchal, Pavel; Struhárová, Iva; Budínská, Eva; Šedo, Ondřej; Vyhlídalová, Tereza; Zdráhal, Zbyněk; Spanning, Rob J.M. van; Kučera, Igor

doi:10.1016/j.bbapap.2010.01.016

Cited by 30 publications

(41 citation statements)

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“…This was independently confirmed at transcript and enzyme activity level (Bouchal et al 2011). Furthermore, synthesis of Fe/Mn superoxide dismutase is independent of FNR-type regulators (Bouchal et al 2010). The only protein in membrane fraction induced synergically by nitrate and azide (spot 1702 in membrane fraction map) was TonB dependent receptor, a protein involved in iron transport.…”

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confidence: 60%

“…Although their level is probably too low for detection on 2-D PAGE gels, an UspA gene product (gene 1849) as a direct neighbour of fnrP in the genome was detected in spot 3211 (Sypro Ruby 2-D map). Its expression profile was similar to fnrP gene product, suggesting their co-expression in single operone (Bouchal et al 2010). Terminal oxidases (aa 3 , cbb 3 and ba 3 types in P. denitrificans, Fig.…”

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confidence: 85%

“…Subsequently, several comprehensive proteomic experiments were performed during the years using our proteomic platform in order to study the differences in protein composition caused by the growth on different terminal electron acceptors in both total cell lysates and membrane fractions ). An additional large proteomic study with data confirmation at transcript level was performed to describe the regulons of three FNR-type transcription regulators FnrP, NNR and NarR at protein level (Bouchal et al 2010). Quantitative and statistical image analysis primarily resulted in creation of local database files in a PDQUEST format.…”

Section: P Denitrificans Proteome Analysis: Methods Developmentmentioning

confidence: 99%

“…The Spearman correlated clustering of the regulation profiles of proteins differentially regulated in response to mutation in FnrP, NNR and NarR transcription regulators and/or to oxygen and nitrate. See (Bouchal et al 2010) for details.…”

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confidence: 99%

“…We also found glyceraldehyde-3-phosphate dehydrogenase (spot 7402 in Coommassie-stained total cell lysate map) non-affected with different growth conditions, so it could serve as an internal standard for gene expression comparison. Our subsequent comprehensive study focused on FNR-type transcription regulators (Bouchal et al 2010) revealed four significant protein clusters according to correlation of their levels under aerobic, semiaerobic and semiaerobic with nitrate growth conditions (see Fig. 8, spot numbers correspond to Sypro Ruby-stained proteome map): (i) The first cluster contains proteins involved in the FnrP regulon.…”

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confidence: 99%

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2D-PAGE Database for Studies on Energetic Metabolism of the Denitrifying Bacterium Paracoccus denitrificans

Bouchal¹,

Stein²,

Zdráhal³

et al. 2012

Integrative Proteomics

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confidence: 60%

mentioning

confidence: 85%

Section: P Denitrificans Proteome Analysis: Methods Developmentmentioning

confidence: 99%

mentioning

confidence: 99%

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confidence: 99%

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2D-PAGE Database for Studies on Energetic Metabolism of the Denitrifying Bacterium Paracoccus denitrificans

Bouchal¹,

Stein²,

Zdráhal³

et al. 2012

Integrative Proteomics

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Biochemical properties of Paracoccus denitrificans FnrP: reactions with molecular oxygen and nitric oxide

et al. 2016

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In Paracoccus denitrificans, three CRP/FNR family regulatory proteins, NarR, NnrR and FnrP, control the switch between aerobic and anaerobic (denitrification) respiration. FnrP is a [4Fe–4S] cluster-containing homologue of the archetypal O2 sensor FNR from E. coli and accordingly regulates genes encoding aerobic and anaerobic respiratory enzymes in response to O2, and also NO, availability. Here we show that FnrP undergoes O2-driven [4Fe–4S] to [2Fe–2S] cluster conversion that involves up to 2 O2 per cluster, with significant oxidation of released cluster sulfide to sulfane observed at higher O2 concentrations. The rate of the cluster reaction was found to be ~sixfold lower than that of E. coli FNR, suggesting that FnrP can remain transcriptionally active under microaerobic conditions. This is consistent with a role for FnrP in activating expression of the high O2 affinity cytochrome c oxidase under microaerobic conditions. Cluster conversion resulted in dissociation of the transcriptionally active FnrP dimer into monomers. Therefore, along with E. coli FNR, FnrP belongs to the subset of FNR proteins in which cluster type is correlated with association state. Interestingly, two key charged residues, Arg140 and Asp154, that have been shown to play key roles in the monomer–dimer equilibrium in E. coli FNR are not conserved in FnrP, indicating that different protomer interactions are important for this equilibrium. Finally, the FnrP [4Fe–4S] cluster is shown to undergo reaction with multiple NO molecules, resulting in iron nitrosyl species and dissociation into monomers.Electronic supplementary materialThe online version of this article (doi:10.1007/s00775-015-1326-7) contains supplementary material, which is available to authorized users.

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Kinetics of anaerobic elemental sulfur oxidation by ferric iron in Acidithiobacillus ferrooxidans and protein identification by comparative 2-DE-MS/MS

Kučera

Bouchal

Cerná

et al. 2011

Antonie van Leeuwenhoek

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Elemental sulfur oxidation by ferric iron in Acidithiobacillus ferrooxidans was investigated. The apparent Michaelis constant for ferric iron was 18.6 mM. An absence of anaerobic ferric iron reduction ability was observed in bacteria maintained on elemental sulfur for an extended period of time. Upon transition from ferrous iron to elemental sulfur medium, the cells exhibited similar kinetic characteristics of ferric iron reduction under anaerobic conditions to those of cells that were originally maintained on ferrous iron. Nevertheless, a total loss of anaerobic ferric iron reduction ability after the sixth passage in elemental sulfur medium was demonstrated. The first proteomic screening of total cell lysates of anaerobically incubated bacteria resulted in the detection of 1599 protein spots in the master two-dimensional electrophoresis gel. A set of 59 more abundant and 49 less abundant protein spots that changed their protein abundances in an anaerobiosis-dependent manner was identified and compared to iron- and sulfur-grown cells, respectively. Proteomic analysis detected a significant increase in abundance under anoxic conditions of electron transporters, such as rusticyanin and cytochrome c(552), involved in the ferrous iron oxidation pathway. Therefore we suggest the incorporation of rus-operon encoded proteins in the anaerobic respiration pathway. Two sulfur metabolism proteins were identified, pyridine nucleotide-disulfide oxidoreductase and sulfide-quinone reductase. The important transcription regulator, ferric uptake regulation protein, was anaerobically more abundant. The anaerobic expression of several proteins involved in cell envelope formation indicated a gradual adaptation to elemental sulfur oxidation.

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Unraveling an FNR based regulatory circuit in Paracoccus denitrificans using a proteomics-based approach

Cited by 30 publications

References 29 publications

2D-PAGE Database for Studies on Energetic Metabolism of the Denitrifying Bacterium Paracoccus denitrificans

2D-PAGE Database for Studies on Energetic Metabolism of the Denitrifying Bacterium Paracoccus denitrificans

Biochemical properties of Paracoccus denitrificans FnrP: reactions with molecular oxygen and nitric oxide

Kinetics of anaerobic elemental sulfur oxidation by ferric iron in Acidithiobacillus ferrooxidans and protein identification by comparative 2-DE-MS/MS

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