scholarly journals Novel nitrite reductase domain structure suggests a chimeric denitrification repertoire in Phylum Chloroflexi

2021 ◽  
Author(s):  
Sarah Schwartz ◽  
Lily Momper ◽  
L. Thiberio Rangel ◽  
Cara Magnabosco ◽  
Jan Amend ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Nitrite Reductase ◽  
Phylogenetic Analyses ◽  
Domain Architecture ◽  
Terrestrial Ecosystems ◽  
Genomic Diversity ◽  
Nitric Oxide Reductase ◽  
Fusion Event ◽  
Ecological Carrying Capacity ◽  
Global Nitrogen Cycle

Denitrification plays a central role in the global nitrogen cycle, reducing and removing nitrogen from marine and terrestrial ecosystems. The flux of nitrogen species through this pathway has a widespread impact, affecting ecological carrying capacity, agriculture, and climate. Nitrite reductase (Nir) and nitric oxide reductase (NOR) are the two central enzymes in this pathway. Here we present a previously unreported Nir domain architecture in members of Phylum Chloroflexi. Phylogenetic analyses of protein domains within Nir indicate that an ancestral horizontal transfer and fusion event produced this chimeric domain architecture. We also identify an expanded genomic diversity of a rarely reported nitric oxide reductase subtype, eNOR. Together, these results suggest a greater diversity of denitrification enzyme arrangements exist than have been previously reported.

scholarly journals Functional interactions between nitrite reductase and nitric oxide reductase from Paracoccus denitrificans

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Ingrid Albertsson ◽  
Johannes Sjöholm ◽  
Josy ter Beek ◽  
Nicholas J. Watmough ◽  
Jerker Widengren ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Nitrite Reductase ◽  
Paracoccus Denitrificans ◽  
Affinity Constant ◽  
Nitric Oxide Reductase ◽  
Terminal Electron Acceptor ◽  
Model Soil ◽  
Transient Interactions ◽  
Electron Donation

AbstractDenitrification is a microbial pathway that constitutes an important part of the nitrogen cycle on earth. Denitrifying organisms use nitrate as a terminal electron acceptor and reduce it stepwise to nitrogen gas, a process that produces the toxic nitric oxide (NO) molecule as an intermediate. In this work, we have investigated the possible functional interaction between the enzyme that produces NO; the cd1 nitrite reductase (cd1NiR) and the enzyme that reduces NO; the c-type nitric oxide reductase (cNOR), from the model soil bacterium P. denitrificans. Such an interaction was observed previously between purified components from P. aeruginosa and could help channeling the NO (directly from the site of formation to the side of reduction), in order to protect the cell from this toxic intermediate. We find that electron donation to cNOR is inhibited in the presence of cd1NiR, presumably because cd1NiR binds cNOR at the same location as the electron donor. We further find that the presence of cNOR influences the dimerization of cd1NiR. Overall, although we find no evidence for a high-affinity, constant interaction between the two enzymes, our data supports transient interactions between cd1NiR and cNOR that influence enzymatic properties of cNOR and oligomerization properties of cd1NiR. We speculate that this could be of particular importance in vivo during metabolic switches between aerobic and denitrifying conditions.

scholarly journals Identification, Functional Studies, and Genomic Comparisons of New Members of the NnrR Regulon in Rhodobacter sphaeroides

2009 ◽  
Vol 192 (4) ◽  
pp. 903-911 ◽  
Author(s):  
Angela Hartsock ◽  
James P. Shapleigh
Keyword(s):  
Nitric Oxide ◽  
Rhodobacter Sphaeroides ◽  
Nitrite Reductase ◽  
Reductase Activity ◽  
Nitrite Reduction ◽  
No Production ◽  
Nitric Oxide Reductase ◽  
New Members ◽  
Functional Studies ◽  
The Status

ABSTRACT Analysis of the Rhodobacter sphaeroides 2.4.3 genome revealed four previously unidentified sequences similar to the binding site of the transcriptional regulator NnrR. Expression studies demonstrated that three of these sequences are within the promoters of genes, designated paz, norEF, and cdgA, in the NnrR regulon, while the status of the fourth sequence, within the tat operon promoter, remains uncertain. nnrV, under control of a previously identified NnrR site, was also identified. paz encodes a pseudoazurin that is a donor of electrons to nitrite reductase. paz inactivation did not decrease nitrite reductase activity, but loss of pseudoazurin and cytochrome c2 together reduced nitrite reduction. Inactivation of norEF reduced nitrite and nitric oxide reductase activity and increased the sensitivity to nitrite in a taxis assay. This suggests that loss of norEF increases NO production as a result of decreased nitric oxide reductase activity. 2.4.3 is the only strain of R. sphaeroides with norEF, even though all four of the strains whose genomes have been sequenced have the norCBQD operon and nnrR. norEF was shown to provide resistance to nitrite when it was mobilized into R. sphaeroides strain 2.4.1 containing nirK. Inactivation of the other identified genes did not reveal any detectable denitrification-related phenotype. The distribution of members of the NnrR regulon in R. sphaeroides revealed patterns of coselection of structural genes with the ancillary genes identified here. The strong coselection of these genes indicates their functional importance under real-world conditions, even though inactivation of the majority of them does not impact denitrification under laboratory conditions.

Separation of soluble denitrifying enzymes and cytochromes from Pseudomonas perfectomarinus

1973 ◽  
Vol 19 (7) ◽  
pp. 861-872 ◽  
Author(s):  
C. D. Cox Jr. ◽  
W. J. Payne
Keyword(s):  
Nitric Oxide ◽  
Electron Donor ◽  
Nitrite Reductase ◽  
Reductase Activity ◽  
Electron Flow ◽  
Deae Cellulose ◽  
The Other ◽  
Nitric Oxide Reductase ◽  
Effective Electron ◽  
Nitrite Reductase Activity

Nitrite and nitric oxide reductases were found soluble in extracts of Pseudomonas perfectomarinus cultured anaerobically at the expense of nitrate and ruptured with the French pressure cell. Malic enzyme, transhydrogenase, and flavin reductase that provided electron flow for these reductases were soluble as well. Nitrous oxide reductase remained particle-bound. Exogenous NADH was a poor electron donor for crude extracts, but a combination of malate, NADP, and NAD served well in the reduction of nitrite and nitric oxide. Nitrite reductase activity lost on dialysis of crude extract was restored by addition of this combination. Addition of free flavins was required for reduction of nitrite and nitric oxide. A nitrite reductase complex was separated from the nitric oxide reductase by gel filtration and DEAE-cellulose chromatography. NADH was an effective electron donor for this system with flavins provided as well. A c-type cytochrome with a split-α peak (perhaps associated with a d type) and two additional c-type cytochromes were separated from the nitrite reductase fraction. One of the latter (RI) emerged oxidized, the other (RII) reduced. Only nitric oxide oxidized RII. When these cytochromes were added to reaction mixtures containing nitrite reductase, activity was increased most by the split-α fraction. After reduction with dithionite, the absorption spectrum of the split-α cytochrome was returned to the oxidized spectrum by addition of nitrite but not the other oxides. A significant amount of a c-type cytochrome remained bound to the nitric oxide reductase fraction. A combination of malic acid, NAD, and NADP was more effective than NADH as electron donor for this system with free flavins provided as well. Addition of RI increased the rate of nitric oxide reduction by this fraction.

scholarly journals Diversity and evolution of nitric oxide reduction

2021 ◽  
Author(s):  
James Hemp ◽  
Ranjani Murali ◽  
Laura A Pace ◽  
Robert A Sanford ◽  
Roland Hatzenpichler ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Active Site ◽  
Phylogenetic Analyses ◽  
Essential Element ◽  
Nitric Oxide Reductase ◽  
Oxide Reduction ◽  
Rhodothermus Marinus ◽  
Fixed Nitrogen ◽  
Proton Channels ◽  
Nitric Oxide Reduction

Nitrogen is an essential element for life, with the availability of fixed nitrogen limiting productivity in many ecosystems. The return of oxidized nitrogen species to the atmospheric N2 pool is predominately catalyzed by microbial denitrification (NO3- → NO2- → NO → N2O → N2). Incomplete denitrification can produce N2O as a terminal product, leading to an increase in atmospheric N2O, a potent greenhouse and ozone depleting gas2. The production of N2O is catalyzed by nitric oxide reductase (NOR) members of the heme-copper oxidoreductase (HCO) superfamily3. Here we propose that a number of uncharacterized HCO families perform nitric oxide reduction and demonstrate that an enzyme from Rhodothermus marinus, belonging to one of these families does perform nitric oxide reduction. These families have novel active-site structures and several have conserved proton channels, suggesting that they might be able to couple nitric oxide reduction to energy conservation. They also exhibit broad phylogenetic and environmental distributions, expanding the diversity of microbes that can perform denitrification. Phylogenetic analyses of the HCO superfamily demonstrate that nitric oxide reductases evolved multiple times independently from oxygen reductases, suggesting that complete denitrification evolved after aerobic respiration.

scholarly journals A QM/MM Study of Nitrite Binding Modes in a Three-Domain Heme-Cu Nitrite Reductase

Molecules ◽  
2018 ◽  
Vol 23 (11) ◽  
pp. 2997
Author(s):  
Kakali Sen ◽  
Michael Hough ◽  
Richard Strange ◽  
Chin Yong ◽  
Thomas Keal
Keyword(s):  
Nitric Oxide ◽  
Nitrite Reductase ◽  
Binding Mode ◽  
Nitrite Reduction ◽  
Quantum Mechanical ◽  
Binding Modes ◽  
Crystallographic Study ◽  
Nitrite Reductases ◽  
Global Nitrogen Cycle ◽  
Oxygen Mode

Copper-containing nitrite reductases (CuNiRs) play a key role in the global nitrogen cycle by reducing nitrite (NO2−) to nitric oxide, a reaction that involves one electron and two protons. In typical two-domain CuNiRs, the electron is acquired from an external electron-donating partner. The recently characterised Rastonia picketti (RpNiR) system is a three-domain CuNiR, where the cupredoxin domain is tethered to a heme c domain that can function as the electron donor. The nitrite reduction starts with the binding of NO2− to the T2Cu centre, but very little is known about how NO2− binds to native RpNiR. A recent crystallographic study of an RpNiR mutant suggests that NO2− may bind via nitrogen rather than through the bidentate oxygen mode typically observed in two-domain CuNiRs. In this work we have used combined quantum mechanical/molecular mechanical (QM/MM) methods to model the binding mode of NO2− with native RpNiR in order to determine whether the N-bound or O-bound orientation is preferred. Our results indicate that binding via nitrogen or oxygen is possible for the oxidised Cu(II) state of the T2Cu centre, but in the reduced Cu(I) state the N-binding mode is energetically preferred.

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