Characterization of a novel Cu-containing dissimilatory nitrite reductase from the haloarchaeon Halorussus sp. YCN54

Dissimilatory nitrite reductase, a key enzyme in the denitrification pathway, catalyzes the reduction of nitrite to NO. Bioinformatic analysis showed that the genome of a novel nitrite-degrading haloarchaeon Halorussus sp. YCN54 possessed a gene encoding the Cu-containing dissimilatory nitrite reductase (NirK(Hrs)). NirK(Hrs) was heterologously expressed and purified. Protein sequencing indicated that two isoforms of NirK(Hrs) monomer were produced intracellularly. UV-vis spectrum of the purified NirK(Hrs) showed that it belonged to the blue NirK group. NirK(Hrs) showed optimum activity at 4.5 M NaCl, 55 celcius and pH 7.0, representing a halophilic, slightly thermophilic and neutral enzyme. It exhibited high stability at 30-50 celcius. NirK(Hrs) activity was strongly inhibited by the copper chelating agent due to removal of copper. NirK(Hrs) activity was activated by Mn2+ and Sr2+. It displayed good tolerance to some high polarity organic solvents and nonionic surfactants, such as glycerol, DMSO, DMF and tween-20. Na2S2O4 was an effective electron donor to NirK(Hrs). The K-m and V-max values of purified NirK(Hrs) for nitrite were 3.2 mM and 477.2 U/mg, respectively, indicating its high activity. These results indicated that NirK(Hrs) may have potential applications for nitrite degradation in high-salt industries, such as salted food and saline wastewater treatment.