Unraveling the mechanism of assimilatory nitrate reduction and methane oxidation by Methylobacter sp. YHQ through dual N-O isotope analysis and kinetic modeling

Abstract

Assimilatory nitrate reduction and methane (CH4) oxidation by bacteria play important roles in carbon (C) and nitrogen (N) biogeochemical cycles. Here, an investigation of enzymatic assimilatory nitrate reduction and CH4 oxidation by Methylobacter sp. YHQ from the wetlands is presented, specifically concentrating on N and oxygen (O) isotope fractionation with various initial nitrate and oxygen concentrations. The N enrichment factors (15εassimilation) increased from 4.2 ± 0.7‰ to 6.9 ±1.3‰ and the O isotope enrichment factors (18εassimilation) increased from 2.7 ± 0.9‰ to 4.7 ± 0.8‰ during nitrate assimilation when initial nitrate concentrations increased from 0.9 mM to 2 mM. Similar 18ε and 15ε values were observed at different oxygen concentrations. The values of 18ε and 15ε provided vital parameters for the assessment of assimilatory nitrate reduction via the Rayleigh equation approach. The ratios of O and N isotope enrichment factors (18ε:15ε)assimilation ranged from 0.64 ± 0.15 to 0.74 ± 0.18 during nitrate assimilation by Methylobacter sp. YHQ with Nas, which were different from (18ε:15ε)assimilation for assimilatory eukaryotic nitrate reductase (eukNR) from literature data. Thus, N and O isotope fractionation could be useful tools to distinguish eukNR from Nas during nitrate assimilation. Additionally, the rates of CH4 oxidation and nitrate reduction were evaluated with a reaction-based kinetic model, and it quantitatively described the enzymatic reactions of nitrate assimilation. Combining dual N-O isotope analysis with kinetic modeling provides new insights into the microbially driven C-N interactions.Graphical