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Mooshammer, Maria; Alves, Ricardo J. E.; Bayer, Barbara; Melcher, Michael; Stieglmeier, Michaela; Jochum, Lara; Rittmann, Simon K-M R.; Watzka, Margarete; Schleper, Christa; Herndl, Gerhard J.; Wanek, Wolfgang (28. July 2020): Nitrogen Isotope Fractionation During Archaeal Ammonia Oxidation: Coupled Estimates From Measurements of Residual Ammonium and Accumulated Nitrite. In: Frontiers in Microbiology, Vol. 11, 1710: pp. 1-9
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Abstract

The naturally occurring nitrogen (N) isotopes,N-15 and(14)N, exhibit different reaction rates during many microbial N transformation processes, which results in N isotope fractionation. Such isotope effects are critical parameters for interpreting natural stable isotope abundances as proxies for biological process rates in the environment across scales. The kinetic isotope effect of ammonia oxidation (AO) to nitrite (NO2-), performed by ammonia-oxidizing archaea (AOA) and ammonia-oxidizing bacteria (AOB), is generally ascribed to the enzyme ammonia monooxygenase (AMO), which catalyzes the first step in this process. However, the kinetic isotope effect of AMO, or epsilon(AMO), has been typically determined based on isotope kinetics during product formation (cumulative product, NO2-) alone, which may have overestimated epsilon(AMO)due to possible accumulation of chemical intermediates and alternative sinks of ammonia/ammonium (NH3/NH4+). Here, we analyzed(15)N isotope fractionation during archaeal ammonia oxidation based on both isotopic changes in residual substrate (RS, NH4+) and cumulative product (CP, NO2-) pools in pure cultures of the soil strainNitrososphaera viennensisEN76 and in highly enriched cultures of the marine strainNitrosopumilus adriaticusNF5, under non-limiting substrate conditions. We obtained epsilon(AMO)values of 31.9-33.1 parts per thousand for both strains based on RS (delta(NH4+)-N-15) and showed that estimates based on CP (delta(NO2-)-N-15) give larger isotope fractionation factors by 6-8 parts per thousand. Complementary analyses showed that, at the end of the growth period, microbial biomass was(15)N-enriched (10.1 parts per thousand), whereas nitrous oxide (N2O) was highly(15)N depleted (-38.1 parts per thousand) relative to the initial substrate. Although we did not determine the isotope effect of NH(4)(+)assimilation (biomass formation) and N2O production by AOA, our results nevertheless show that the discrepancy between epsilon(AMO)estimates based on RS and CP might have derived from the incorporation of(15)N-enriched residual NH(4)(+)after AMO reaction into microbial biomass and that N2O production did not affect isotope fractionation estimates significantly.