Immunogold Localization of Key Metabolic Enzymes in the Anammoxosome and on the Tubule-Like Structures of Kuenenia stuttgartiensis

Almeida, Naomi M. de; Neumann, Sarah; Mesman, Rob; Ferousi, Christina; Keltjens, Jan T.; Jetten, Mike S. M.; Kartal, Boran; Niftrik, Laura van

doi:10.1128/jb.00186-15

Cited by 55 publications

(35 citation statements)

References 37 publications

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“…Thus, the differences in isotope effects may reflect the NxrA phylogeny. However, anammox-bacteria oxidize nitrite in the anammoxosome (de Almeida et al 2015) with an inverse isotope effect of about 31 ‰ (Brunner et al 2013), which is much more pronounced than what we found in Nitrospina and Nitrospira. Indeed, the previously determined isotope effects of Nitro bacter and Nitrococcus are apparently more pronounced and thus closer to that of anammox bacteria.…”

Section: Inverse Kinetic Isotope Effects During Nitrite Oxidationcontrasting

confidence: 52%

Oxidation kinetics and inverse isotope effect of marine nitrite-oxidizing isolates

Jacob¹,

Nowka²,

Merten³

et al. 2017

Aquat. Microb. Ecol.

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Nitrification, the step-wise oxidation of ammonium to nitrite and nitrate, is important in the marine environment because it produces nitrate, the most abundant marine dissolved inorganic nitrogen (DIN) component and N-source for phytoplankton and microbes. This study focused on the second step of nitrification, which is carried out by a distinct group of organisms, nitrite-oxidizing bacteria (NOB). The growth of NOB is characterized by nitrite oxidation kinetics, which we investigated for 4 pure cultures of marine NOB (Nitrospina watsonii 347, Nitrospira sp. Ecomares 2.1, Nitrococcus mobilis 231, and Nitrobacter sp. 311). We further compared the kinetics to those of non-marine species because substrate concentrations in marine environments are comparatively low, which likely influences kinetics and highlights the importance of this study. We also determined the isotope effect during nitrite oxidation of a pure culture of Nitrospina (Nitrospina watsonii 347) belonging to one of the most abundant marine NOB genera, and for a Nitrospira strain (Nitrospira sp. Ecomares 2.1). The enzyme kinetics of nitrite oxidation, described by Michaelis-Menten kinetics, of 4 marine genera are rather narrow and fall in the low end of halfsaturation constant (K m ) values reported so far, which span over 3 orders of magnitude between 9 and >1000 μM NO 2 − . Nitrospina has the lowest K m (19 μM NO 2 − ), followed by Nitrobacter (28 μM NO 2 − ), Nitrospira (54 μM NO 2 − ), and Nitrococcus (120 μM NO 2 − ). The isotope effects during nitrite oxidation by Nitrospina watsonii 347 and Nitrospira sp. Ecomares 2.1 were 9.7 ± 0.8 and 10.2 ± 0.9 ‰, respectively. This confirms the inverse isotope effect of NOB described in other studies; however, it is at the lower end of reported isotope effects. We speculate that differences in isotope effects reflect distinct nitrite oxidoreductase (NXR) enzyme orientations.

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Section: Inverse Kinetic Isotope Effects During Nitrite Oxidationcontrasting

confidence: 52%

Oxidation kinetics and inverse isotope effect of marine nitrite-oxidizing isolates

Jacob¹,

Nowka²,

Merten³

et al. 2017

Aquat. Microb. Ecol.

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“…S1). Cleavage of the N-terminal leader sequence (32 amino acids) would facilitate export of HDH into the anammoxosome, an anammox-specific cell organelle where catabolism resides and where processed HDH is specifically localized as recently shown by immunogold labelling (16).…”

Section: Resultsmentioning

confidence: 94%

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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107

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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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“…S9). Based on the known cell biology ( 64 ), biochemistry and anammox metabolism ( 4, 65, 66 ), and the expression profiles of the known anammox pathways obtained with the differential expression analysis done in this study (Table S3 and 4), we propose a putative molecular model to describe how electrons flow from the anammoxosome to the electrode in the EET-dependent anammox process (Fig. S10).…”

Section: Supplementary Materialsmentioning

confidence: 99%

Extracellular electron transfer-dependent anaerobic oxidation of ammonium by anammox bacteria

Shaw

Ali

Katuri

et al. 2019

Preprint

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AbstractAnaerobic ammonium oxidation (anammox) by anammox bacteria contributes significantly to the global nitrogen cycle, and plays a major role in sustainable wastewater treatment. Anammox bacteria convert ammonium (NH4+) to dinitrogen gas (N2) using nitrite (NO2−) or nitric oxide (NO) as the electron acceptor. In the absence of NO2− or NO, anammox bacteria can couple formate oxidation to the reduction of metal oxides such as Fe(III) or Mn(IV). Their genomes contain homologs of Geobacter and Shewanella cytochromes involved in extracellular electron transfer (EET). However, it is still unknown whether anammox bacteria have EET capability and can couple the oxidation of NH4+ with transfer of electrons to carbon-based insoluble extracellular electron acceptors. Here we show using complementary approaches that in the absence of NO2−, freshwater and marine anammox bacteria couple the oxidation of NH4+ with transfer of electrons to carbon-based insoluble extracellular electron acceptors such as graphene oxide (GO) or electrodes poised at a certain potential in microbial electrolysis cells (MECs). Metagenomics, fluorescence in-situ hybridization and electrochemical analyses coupled with MEC performance confirmed that anammox electrode biofilms were responsible for current generation through EET-dependent oxidation of NH4+. 15N-labelling experiments revealed the molecular mechanism of the EET-dependent anammox process. NH4+ was oxidized to N2 via hydroxylamine (NH2OH) as intermediate when electrode was the terminal electron acceptor. Comparative transcriptomics analysis supported isotope labelling experiments and revealed an alternative pathway for NH4+ oxidation coupled to EET when electrode is used as electron acceptor compared to NO2−as electron acceptor. To our knowledge, our results provide the first experimental evidence that marine and freshwater anammox bacteria can couple NH4+ oxidation with EET, which is a significant finding, and challenges our perception of a key player of anaerobic oxidation of NH4+ in natural environments and engineered systems.

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Immunogold Localization of Key Metabolic Enzymes in the Anammoxosome and on the Tubule-Like Structures of Kuenenia stuttgartiensis

Cited by 55 publications

References 37 publications

Oxidation kinetics and inverse isotope effect of marine nitrite-oxidizing isolates

Oxidation kinetics and inverse isotope effect of marine nitrite-oxidizing isolates

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

Extracellular electron transfer-dependent anaerobic oxidation of ammonium by anammox bacteria

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