Alternative nitrogenase activity in the environment and nitrogen cycle implications
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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.
KeywordsNitrogen fixation Alternative nitrogenase Trace metals Nitrogen cycle Stable isotopes
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.
- Bellenger J-P, Wichard T, Xu Y, Kraepiel AML (2011) Essential metals for nitrogen fixation in a free-living N2-fixing bacterium: chelation, homeostasis and high use efficiency. Environ Microbiol 13:1395–1411Google Scholar
- Capone DG (1988) Benthic nitrogen fixation. In: Blackburn TH, Sørensen J (eds) Nitrogen cycling in coastal marine environments. Springer, New YorkGoogle Scholar
- Chisnell JR, Premakumar R, Bishop PE (1988) Purification of a second alternative nitrogenase from a nifhdk deletion strain of Azotobacter vinelandii. J Bacteriol 170:27–33Google Scholar
- Hayes JM (2004) An introduction to isotopic calculation. Oceanographic Institution, Woods HoleGoogle Scholar
- Joerger RD, Loveless TM, Pau RN et al (1990) Nucleotide sequence and mutational analysis of the structural genes for nitrogenase 2 of Azotobacter vinelandii. J Bacteriol 172:3400–3408Google Scholar
- Kaminski P, Batut J, Boistard P (1998) A survey of symbiotic nitrogen fixation by Rhizobia. In: Spaink HP, Kondorosi A, Hooykaas PJJ (eds) The Rhizobiaceae. Springer, NetherlandsGoogle Scholar
- Perry DA, Oren R, Hart SC (2013) Forest ecosystems. JHU Press, BaltimoreGoogle Scholar
- Thiel T (1993) Characterization of genes for an alternative nitrogenase in the cyanobacterium Anabaena variabilis. J Bacteriol 175:6276–6286Google Scholar
- Walmsley J, Kennedy C (1991) Temperature-dependent regulation by molybdenum and vanadium of expression of the structural genes encoding 3 nitrogenases in Azotobacter vinelandii. Appl Environ Microbiol 57:622–624Google Scholar