Biological nitrogen fixation, the main natural input of fixed nitrogen into the biosphere, is catalyzed by Mo-, V-, or Fe-only nitrogenase metalloenzymes. Although “alternative” V- and Fe-only nitrogenase genes are found in many environments, the contribution of these isoenzymes to N2 fixation is unknown. Here we present a new method (ISARA, isotopic acetylene reduction assay) that distinguishes canonical Mo and alternative nitrogenase activities based on in vivo 13C fractionation of acetylene reduction to ethylene (13εMo = 13.1–14.7 ‰, 13εV = 7.5–8.8 ‰, 13εFe = 5.8–6.5 ‰). ISARA analyses indicate significant contributions of alternative nitrogen fixation in boreal cyanolichens and salt marshes (~10–40 % acetylene reduction, ~20–55 % N2 fixed). These results affect the quantitative interpretation of natural abundance 15N data or traditional acetylene reduction assays. They also invite a reexamination of the conditions under which the different nitrogenase isozymes are active and suggest significant interactions between the cycles of nitrogen and trace metals.
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We gratefully acknowledge the Princeton Environmental Institute (Grand Challenge Program), Andlinger Center for Energy and the Environment (Princeton U.), NSF Geobiology (GG-1024553), and the Canadian Research Chair in Terrestrial Biogeochemistry (CRC-950-219383) for funding; M. Saito, J. Waterbury, and I. Valiela (WHOI); Socièté des Espaces de Plein Air (Québec); T. Thiel (U. Missouri); S. Oleynik (Princeton U.); N. Van Oostende (Princeton U.) for their aid in field investigations, the provision of bacterial cultures, or advice on isotope analyses. The authors declare no conflict of interest.
A.M.L.K. and X.Z. designed the experiments; X.Z., D.L.M., A.M.L.K, R.D. performed the experiments, X.Z., D.L.M., R.D. analyzed the data; J-P.B. contributed materials; X.Z, D.L.M., F.M.M. and A.M.L.K co-wrote the paper.
Responsible Editor: Stuart Grandy.
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Zhang, X., McRose, D.L., Darnajoux, R. et al. Alternative nitrogenase activity in the environment and nitrogen cycle implications. Biogeochemistry 127, 189–198 (2016). https://doi.org/10.1007/s10533-016-0188-6
- Nitrogen fixation
- Alternative nitrogenase
- Trace metals
- Nitrogen cycle
- Stable isotopes