Skip to main content
NCBI home page
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1982 Oct;79(19):5871–5875. doi: 10.1073/pnas.79.19.5871

18O isotope shift in 15N NMR analysis of biological N-oxidations: H2O-NO2- exchange in the ammonia-oxidizing bacterium Nitrosomonas

Kristoffer K Andersson *, Stephen B Philson †, Alan B Hooper *,‡
PMCID: PMC347012  PMID: 16593233

Abstract

The 18O/16O shifts in 15N NMR were determined for nitrite (0.13 ppm or 4.2 Hz at 7.05 T) and nitrate (0.056 ppm or 1.7 Hz at 7.05 T) at neutral pH. The technique, which allows clear differentiation between 16O and 18O derivatives of 15N, was used to assess the source of oxygens in nitrite produced by oxidation of ammonia in Nitrosomonas. The two oxygens of nitrite produced by cell-catalyzed oxidation of ammonia or hydroxylamine had the 16O/18O isotope composition of water. Nitrosomonas is shown to catalyze the rapid exchange of oxygen between nitrite and water. The exchange reaction required the concomitant oxidation of ammonia. The amount of nitrite exchanged could exceed the amount of ammonia oxidized by a factor of 3. This exchange explains previous difficulties in the determination of the source of nitrite oxygen in ammonia oxidation. When cells oxidized [15N]ammonia in the presence of a great excess of exogenous [14N]nitrite, 20% of one oxygen in the resulting [15N]nitrite was derived from dioxygen. Dioxygen is apparently the source of at least one oxygen in nitrite produced by Nitrosomonas.

Keywords: hydroxylamine, nitrite, oxygen exchange

Full text

PDF
5871

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Cohn M., Hu A. Isotopic (18O) shift in 31P nuclear magnetic resonance applied to a study of enzyme-catalyzed phosphate--phosphate exchange and phosphate (oxygen)--water exchange reactions. Proc Natl Acad Sci U S A. 1978 Jan;75(1):200–203. doi: 10.1073/pnas.75.1.200. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Dua R. D., Bhandari B., Nicholas D. J. Stable isotope studies on the oxidation of ammonia to hydroxylamine by Nitrosomonas europaea. FEBS Lett. 1979 Oct 15;106(2):401–404. doi: 10.1016/0014-5793(79)80541-4. [DOI] [PubMed] [Google Scholar]
  3. Erickson R. H., Hooper A. B., Terry K. R. Solubilization and purification of cytochrome 1 from Nitrosomonas. Biochim Biophys Acta. 1972;283(1):155–166. doi: 10.1016/0005-2728(72)90107-7. [DOI] [PubMed] [Google Scholar]
  4. Hollocher T. C., Tate M. E., Nicholas D. J. Oxidation of ammonia by Nitrosomonas europaea. Definite 18O-tracer evidence that hydroxylamine formation involves a monooxygenase. J Biol Chem. 1981 Nov 10;256(21):10834–10836. [PubMed] [Google Scholar]
  5. Hooper A. B. A nitrite-reducing enzyme from Nitrosomonas europaea. Preliminary characterization with hydroxylamine ad electron donor. Biochim Biophys Acta. 1968 Jul 16;162(1):49–65. doi: 10.1016/0005-2728(68)90213-2. [DOI] [PubMed] [Google Scholar]
  6. Hooper A. B., Maxwell P. C., Terry K. R. Hydroxylamine oxidoreductase from Nitrosomonas: absorption spectra and content of heme and metal. Biochemistry. 1978 Jul 25;17(15):2984–2989. doi: 10.1021/bi00608a007. [DOI] [PubMed] [Google Scholar]
  7. Hooper A. B., Terry K. R., Maxwell P. C. Hydroxylamine oxidoreductase of Nitrosomonas. Oxidation of diethyldithiocarbamate concomitant with stimulation of nitrite synthesis. Biochim Biophys Acta. 1977 Oct 12;462(1):141–152. doi: 10.1016/0005-2728(77)90196-7. [DOI] [PubMed] [Google Scholar]
  8. Rees M., Nason A. Incorporation of atmospheric oxygen in nitriteformed during ammonia oxidation by Nitrosomonas europaea. Biochim Biophys Acta. 1966 Feb 14;113(2):398–401. [PubMed] [Google Scholar]
  9. Tsai M. D., Huang S. L., Kozlowski J. F., Chang C. C. Applicability of the phosphorus-31 (oxygen-17) nuclear magnetic resonance method in the study of enzyme mechanism involving phosphorus. Biochemistry. 1980 Jul 22;19(15):3531–3536. doi: 10.1021/bi00556a018. [DOI] [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES