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

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.