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Molecular mechanism of anaerobic ammonium oxidation

Nature volume 479, pages 127–130 (2011)Cite this article

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Abstract

Two distinct microbial processes, denitrification and anaerobic ammonium oxidation (anammox), are responsible for the release of fixed nitrogen as dinitrogen gas (N2) to the atmosphere1,2,3,4. Denitrification has been studied for over 100 years and its intermediates and enzymes are well known5. Even though anammox is a key biogeochemical process of equal importance, its molecular mechanism is unknown, but it was proposed to proceed through hydrazine (N2H4)6,7. Here we show that N2H4 is produced from the anammox substrates ammonium and nitrite and that nitric oxide (NO) is the direct precursor of N2H4. We resolved the genes and proteins central to anammox metabolism and purified the key enzymes that catalyse N2H4 synthesis and its oxidation to N2. These results present a new biochemical reaction forging an N–N bond and fill a lacuna in our understanding of the biochemical synthesis of the N2 in the atmosphere. Furthermore, they reinforce the role of nitric oxide in the evolution of the nitrogen cycle.

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Figure 1: Biochemical pathway and enzymatic machinery of K. stuttgartiensis.
Figure 2: Determination of nitric oxide (NO) as an intermediate.
Figure 3: Hydrazine turnover.
Figure 4: 29 N 2 production by hydrazine synthase complex and kustc1061 from 15 NH 4 + and NO.

Accession codes

Data deposits

The metatranscriptome and peptidome sequences are deposited in Gene Expression Omnibus under accession numbers GSE15408 and PSE111, respectively.

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Acknowledgements

B.K. was supported by a grant (05987) from the Dutch Foundation for Applied Research. W.J.M. was supported by a grant (142161201) from the Darwin Center for Biogeosciences. N.M.d.A. and I.C. were supported by a grant (81802015) from the Netherlands Organization for Scientific Research. M.S. was supported by a VIDI grant from the Netherlands Organization for Scientific Research and a European Research Council grant MASEM (242635). The anammox research of M.S.M.J. is supported by an advanced grant (232987) from the ERC. The authors acknowledge R. Klefoth for the initial tests for protein purification procedures.

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Authors and Affiliations

  1. Department of Microbiology, Institute for Water and Wetland Research, Radboud University Nijmegen, Heyendaalseweg 135, 6525AJ Nijmegen, The Netherlands,

    Boran Kartal, Wouter J. Maalcke, Naomi M. de Almeida, Irina Cirpus, Wim Geerts, Huub J. M. Op den Camp, Harry R. Harhangi, Jan T. Keltjens, Mike S. M. Jetten & Marc Strous

  2. Department of Laboratory Medicine, Nijmegen Centre for Mitochondrial Disorders, Nijmegen Proteomics Facility, Radboud University Nijmegen Medical Centre, Geert Grooteplein 10, 6500HB Nijmegen, The Netherlands,

    Jolein Gloerich

  3. Department of Molecular Biology, Radboud University, Nijmegen Centre for Molecular Life Sciences, Geert Grooteplein 28, 6525 GA Nijmegen, The Netherlands,

    Eva M. Janssen-Megens, Kees-Jan Francoijs & Hendrik G. Stunnenberg

  4. Department of Biotechnology, Delft University of Technology, 2628 BC Delft, The Netherlands,

    Mike S. M. Jetten

  5. Max Planck Institute for Marine Microbiology, Celsiusstrasse 1, 28359 Bremen, Germany ,

    Marc Strous

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Contributions

Physiological experiments were conceived, designed and performed by B.K., Kuenenia stuttgartiensis was grown by B.K. and W.G., two-dimensional gel electrophoresis was performed by N.M.A. and I.C., one-dimensional gel electrophoresis was performed by W.J.M. and B.K., MALDI–TOF analysis was performed by B.K., W.J.M. and H.J.M.O.d.C., nanoLC-MS/MS by J.G., RNA extraction and reverse transcription by H.R.H., Illumina sequencing by E.M.J.-M., K.-J.F. and H.S., and protein purification and activity tests were designed by W.J.M., B.K. and J.T.K. and performed by W.J.M. Proteomic and transcriptomic data processing was performed by J.G., M.S., K.-J.F., B.K., M.S.M.J. and H.J.M.O.d.C. The manuscript was written by B.K. with input from J.T.K., M.S. and M.S.M.J.

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Correspondence to Boran Kartal.

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Kartal, B., Maalcke, W., de Almeida, N. et al. Molecular mechanism of anaerobic ammonium oxidation. Nature 479, 127–130 (2011). https://doi.org/10.1038/nature10453

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