The inner workings of the hydrazine synthase multiprotein complex

Dietl, Andreas; Ferousi, Christina; Maalcke, Wouter J.; Menzel, Andreas; Vries, Simon de; Keltjens, Jan T.; Jetten, Mike S. M.; Kartal, Boran; Barends, Thomas R. M.

doi:10.1038/nature15517

Cited by 136 publications

(122 citation statements)

References 34 publications

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“…Octaheme KsHOX from K. stuttgartiensis (kustc1061) and from other anammox bacteria oxidizes hydrazine as well, albeit with lower catalytic efficiency (k cat / K m ). Hydrazine also served as a substrate for NeHAO, surprisingly with k cat and K m values that were superior to genuine HDH, even though hydrazine does not play a role in N. europaea metabolism: this organism and other aerobic ammonium oxidizing microorganisms are not able synthesize hydrazine, which is an exclusive property of anammox bacteria (3,(5)(6)(7).…”

Section: Resultsmentioning

confidence: 99%

Characterization of Anammox Hydrazine Dehydrogenase, a Key N2-producing Enzyme in the Global Nitrogen Cycle

Maalcke

Reimann

Vries

et al. 2016

Journal of Biological Chemistry

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104

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Anaerobic ammonium-oxidizing (anammox) bacteria derive their energy for growth from the oxidation of ammonium with nitrite as the electron acceptor. N 2 , the end product of this metabolism, is produced from the oxidation of the intermediate, hydrazine (N 2 H 4 ). Previously, we identified N 2 -producing hydrazine dehydrogenase (KsHDH) from the anammox organism Kuenenia stuttgartiensis as the gene product of kustc0694 and determined some of its catalytic properties. In the genome of K. stuttgartiensis, kustc0694 is one of 10 paralogs related to octaheme hydroxylamine (NH 2 OH) oxidoreductase (HAO). Here, we characterized KsHDH as a covalently cross-linked homotrimeric octaheme protein as found for HAO and HAO-related hydroxylamine-oxidizing enzyme kustc1061 from K. stuttgartiensis. Interestingly, the HDH trimers formed octamers in solution, each octamer harboring an amazing 192 c-type heme moieties. Whereas HAO and kustc1061 are capable of hydrazine oxidation as well, KsHDH was highly specific for this activity. To understand this specificity, we performed detailed amino acid sequence analyses and investigated the catalytic and spectroscopic (electronic absorbance, EPR) properties of KsHDH in comparison with the well defined HAO and kustc1061. We conclude that HDH specificity is most likely derived from structural changes around the catalytic heme 4 (P 460 ) and of the electron-wiring circuit comprising seven His/His-ligated c-type hemes in each subunit. These nuances make HDH a globally prominent N 2 -producing enzyme, next to nitrous oxide (N 2 O) reductase from denitrifying microorganisms.An estimated 30 -70% of all N 2 that is released into the atmosphere is produced by anaerobic ammonium-oxidizing (anammox) 4 bacteria (1, 2), which represent one of the latest scientific discoveries in the biogeochemical nitrogen cycle. These organisms gain their energy for growth from the oxidation of ammonium, with nitrite as the electron acceptor, to produce N 2 . With the employment of advanced molecular tools, these bacteria have been detected in nearly every anoxic environment where fixed nitrogen compounds are degraded (3). Besides its biogeochemical and ecological relevance, the anammox process has found worldwide application in ammonium removal from wastewater as an environment-friendly and cost-effective alternative to conventional systems (4).In our current understanding, anammox catabolism is composed of three consecutive, coupled reactions with two intermediates, nitric oxide (NO) and hydrazine (N 2 H 4 ): 1) the oneelectron reduction of the substrate nitrite to NO (Reaction 1); 2) the activation of the second substrate ammonium with NO and the concomitant input of three electrons to synthesize N 2 H 4 (Reaction 2); and 3) the oxidation of hydrazine, the most powerful reductant in nature, to N 2 (Reaction 3) (5-7). NO

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Section: Resultsmentioning

confidence: 99%

Characterization of Anammox Hydrazine Dehydrogenase, a Key N2-producing Enzyme in the Global Nitrogen Cycle

Maalcke

Reimann

Vries

et al. 2016

Journal of Biological Chemistry

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104

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“…Anaerobic ammonium oxidation (anammox) is a microbial process in which

{NH}_{4}^{+}

and

{NO}_{2}^{-}

are directly converted to N 2 gas via N 2 H 4 (de Almeida et al ., ; Kartal et al ., ). The proposed biochemical pathway of the anammox process is as follows: first,

{NO}_{2}^{-}

is reduced to nitric oxide (NO) by either cytochrome cd 1 ‐type or copper‐containing nitrite reductase (NirS and NirK, respectively) (Strous et al ., ; Hira et al ., ); then, the produced NO is reduced to hydroxylamine (NH 2 OH) and coupled with NH 3 to form hydrazine (N 2 H 4 ) by hydrazine synthase (Hzs) (Kartal et al ., ; Dietl et al ., ); and finally, the N 2 H 4 is oxidised to N 2 gas by hydrazine dehydrogenase (Hdh) (Shimamura et al ., ; Kartal et al ., ). The anammox process has been demonstrated in bacteria affiliated with the order Brocadiales .…”

Section: Introductionmentioning

confidence: 99%

Hydroxylamine‐dependent anaerobic ammonium oxidation (anammox) by “Candidatus Brocadia sinica”

Oshiki

Ali

Shinyako-Hata³

et al. 2016

Environmental Microbiology

114

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Summary Although metabolic pathways and associated enzymes of anaerobic ammonium oxidation (anammox) of ‘Ca. Kuenenia stuttgartiensis’ have been studied, those of other anammox bacteria are still poorly understood. NO2− reduction to NO is considered to be the first step in the anammox metabolism of ‘Ca. K. stuttgartiensis’, however, ‘Ca. Brocadia’ lacks the genes that encode canonical NO‐forming nitrite reductases (NirS or NirK) in its genome, which is different from ‘Ca. K. stuttgartiensis’. Here, we studied the anammox metabolism of ‘Ca. Brocadia sinica’. 15N‐tracer experiments demonstrated that ‘Ca. B. sinica’ cells could reduce NO2− to NH2OH, instead of NO, with as yet unidentified nitrite reductase(s). Furthermore, N2H4 synthesis, downstream reaction of NO2− reduction, was investigated using a purified ‘Ca. B. sinica' hydrazine synthase (Hzs) and intact cells. Both the ‘Ca. B. sinica’ Hzs and cells utilized NH2OH and NH4+, but not NO and NH4+, for N2H4 synthesis and further oxidized N2H4 to N2 gas. Taken together, the metabolic pathway of ‘Ca. B. sinica’ is NH2OH‐dependent and different from the one of ‘Ca. K. stuttgartiensis’, indicating metabolic diversity of anammox bacteria.

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“…In the case of nitrifying bacteria, several studies hypothesize that the azole compounds bind to the metal center in key metabolic enzymes such as copper in ammonium monooxygenase, which results in toxicity . Similarly, many key enzymes of anammox bacteria contain metal prosthetic groups such as copper in nitrite reductase, and the copper or zinc in hydrazine synthase . Most importantly, anammox bacteria contain very high amounts of cytochrome C, a hemeprotein which contains iron .…”

Section: Discussionmentioning

confidence: 99%

Toxicity of azoles towards the anaerobic ammonium oxidation (anammox) process

Lakhey

Sierra-Álvarez

et al. 2019

J of Chemical Tech & Biotech

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BACKGROUND Azoles are an important class of compounds that are widely used as corrosion inhibitors in aircraft de‐icing agents, cooling towers, semiconductor manufacturing and household dishwashing detergents. They also are important moieties in pharmaceutical drugs and fungicides. Azoles are widespread emerging contaminants occurring frequently in water bodies. Azole compounds can potentially cause inhibition towards key biological processes in natural ecosystems and wastewater treatment processes. Of particular concern is the inhibition of azoles to the nitrification process (aerobic oxidation of ammonium). This study investigated the acute toxicity of azole compounds towards the anaerobic ammonia oxidation (anammox) process, which is an important environmental biotechnology gaining traction for nutrient‐nitrogen removal during wastewater treatment. In this study, using batch bioassay techniques, the anammox toxicity of eight commonly occurring azole compounds was evaluated. RESULTS The results show that 1H‐benzotriazole and 5‐methyl‐1H‐benzotriazole had the highest inhibitory effect on the anammox process, causing 50% decrease in anammox activity (IC50) at concentrations of 19.6 and 17.8 mg L−1, respectively. 1H‐imidazole caused less severe toxicity with an IC50 of 79.4 mg L−1. The other azole compounds were either nontoxic (1H‐pyrazole, 1H‐1,2,4‐triazole and 1‐methyl‐pyrazole) or at best mildly toxic (1H‐benzotriazole‐5‐carboxylic acid and 3,5‐dimethyl‐1H‐pyrazole) towards the anammox bacteria at the concentrations tested. CONCLUSIONS This study showed that most azole compounds tested displayed mild to low or no toxicity towards the anammox bacteria. The anammox bacteria were found to be far less sensitive to azoles compared to nitrifying bacteria. © 2019 Society of Chemical Industry

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The inner workings of the hydrazine synthase multiprotein complex

Cited by 136 publications

References 34 publications

Characterization of Anammox Hydrazine Dehydrogenase, a Key N2-producing Enzyme in the Global Nitrogen Cycle

Characterization of Anammox Hydrazine Dehydrogenase, a Key N2-producing Enzyme in the Global Nitrogen Cycle

Hydroxylamine‐dependent anaerobic ammonium oxidation (anammox) by “Candidatus Brocadia sinica”

Toxicity of azoles towards the anaerobic ammonium oxidation (anammox) process

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