The effect of pH on Marinobacter hydrocarbonoclasticus denitrification pathway and nitrous oxide reductase

Abstract

Increasing atmospheric concentration of N2O has been a concern, as it is a potent greenhouse gas and promotes ozone layer destruction. In the N-cycle, release of N2O is boosted upon a drop of pH in the environment. Here, Marinobacter hydrocarbonoclasticus was grown in batch mode in the presence of nitrate, to study the effect of pH in the denitrification pathway by gene expression profiling, quantification of nitrate and nitrite, and evaluating the ability of whole cells to reduce NO and N2O. At pH 6.5, accumulation of nitrite in the medium occurs and the cells were unable to reduce N2O. In addition, the biochemical properties of N2O reductase isolated from cells grown at pH 6.5, 7.5 and 8.5 were compared for the first time. The amount of this enzyme at acidic pH was lower than that at pH 7.5 and 8.5, pinpointing to a post-transcriptional regulation though pH did not affect gene expression of N2O reductase accessory genes. N2O reductase isolated from cells grown at pH 6.5 has its catalytic center mainly as CuZ(4Cu1S), while that from cells grown at pH 7.5 or 8.5 has it as CuZ(4Cu2S). This study evidences that an in vivo secondary level of regulation is required to maintain N2O reductase in an active state.

Copper availability governs nitrous oxide accumulation in wetland soils and stream sediments

“…Optimal N2O reduction has been observed in the pH range of 7.5-8.0; previous studies have observed substantial N2O accumulation in the pH range of 6.0-6.5 (Pan et al, 2012;Carreira et al, 2020). Although the pH in the riparian wetland soils increased to ~ 6.5 in the first two days of incubation, i.e., before the onset of N2O accumulation, it was in the range where a decrease in the activity of nitrous reductase enzyme has been observed (Pan et al, 2012;Carreira et al, 2020). Thus, Riparian 1 and Riparian 2 wetland soils could be a significant source of N2O, not only because the bioavailable Cu is limited but also because they are acidic.…”

“…One possible explanation is that the riparian wetland incubation experiments were conducted at pH 5, whereas the incubations for marsh wetland soils (Marsh 1) and stream sediments (Stream 1 and Stream 2) were performed at neutral pH conditions. Acidic soils decrease the activity of the nitrous oxide reductase enzyme, leading to N2O accumulation (Knowles, 1982;Simek and Cooper, 2002;Pan et al, 2012;Carreira et al, 2020). Optimal N2O reduction has been observed in the pH range of 7.5-8.0; previous studies have observed substantial N2O accumulation in the pH range of 6.0-6.5 (Pan et al, 2012;Carreira et al, 2020).…”

“…Acidic soils decrease the activity of the nitrous oxide reductase enzyme, leading to N2O accumulation (Knowles, 1982;Simek and Cooper, 2002;Pan et al, 2012;Carreira et al, 2020). Optimal N2O reduction has been observed in the pH range of 7.5-8.0; previous studies have observed substantial N2O accumulation in the pH range of 6.0-6.5 (Pan et al, 2012;Carreira et al, 2020). Although the pH in the riparian wetland soils increased to ~ 6.5 in the first two days of incubation, i.e., before the onset of N2O accumulation, it was in the range where a decrease in the activity of nitrous reductase enzyme has been observed (Pan et al, 2012;Carreira et al, 2020).…”

Effect of pH on the denitrification proteome of the soil bacterium Paracoccus denitrificans PD1222

“…In P. denitrificans , NosZ activity is sensitive to extracellular changes in pH, but nosZ gene expression is not altered, suggesting that pH affects protein synthesis and/or assembly 24 , 25 . Furthermore, the bacterium Marinobacter hydrocarbonoclasticus is unable to reduce N 2 O at pH 6.5, displaying low amounts of copper-loaded NosZ enzyme at this pH 28 . Reduced NosZ activity (Fig.…”

A [3Cu:2S] cluster provides insight into the assembly and function of the CuZ site of nitrous oxide reductase

“… 4,6,10,18,52,53 The H382A variant may now help to reconcile these seemingly incompatible results, suggesting that S Z2 can indeed change its position from ligating Cu 1 of Cu Z to the nearby K454 (similar sulfur-shift mechanism was envisaged by Moura and Pauleta). 53 This would leave both Cu 1 and Cu 4 with three remaining ligands, and thus with the opportunity to bind an additional exogenous ligand, the substrate N 2 O, in a 1,3-bridging fashion. This binding mode is similar to the one proposed by Moura and Solomon, 52 but does not require dissociation of S Z2 in accordance with our structural data.…”