Oxidation kinetics and inverse isotope effect of marine nitrite-oxidizing isolates


Nitrification, the step-wise oxidation of ammonium to nitrite and nitrate, is important in the marine environment, because it produces nitrate, the most abundant marine dissolved inorganic nitrogen (DIN) component and N-source for phytoplankton and microbes. This study focuses on the second step of nitrification, which is carried out by a distinct group of organisms, nitrite-oxidizing bacteria (NOB). The growth of NOB is characterized by nitrite oxidation kinetics, which we investigate for four pure cultures of marine NOB (Nitrospina watsonii 347, Nitrospira Ecomares 2.1, Nitrococcus mobilis 231, and Nitrobacter sp. 311). We further compare the kinetics to those of non-marine species, because substrate concentrations in marine environments are comparatively low, which likely influences kinetics and highlights the importance of this study. We also determine the isotope effect during nitrite oxidation of a pure culture of Nitrospina (Nitrospina watsonii 347), belonging to one of the most abundant marine NOB genera, and for a Nitrospira strain (Nitrospira Ecomares 2.1). The enzyme kinetic of nitrite oxidation, described by Michaelis-Menten kinetics, of four marine genera are rather narrow and fall in the low end of Km values reported so far, which span over three orders of magnitude between 9 to >1000 µM NO2-. Nitrospina has the lowest half-saturation constant Km (19 µM NO2-), followed by Nitrobacter (28 µM NO2-), Nitrospira (Km of 54 µM NO2-), and Nitrococcus (120 µM NO2-). The isotope effect during nitrite oxidation by Nitrospira watsonii 347 and Nitrospina Ecomares 2.1 is 10.2±0.9‰ and 9.7±0.8‰, respectively. This confirms the inverse isotope effect of NOB described previously; however, it is at the lower end of reported isotope effects. We speculate that differences in isotope effects reflect distinct NXR orientation.
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