Purification and biochemical characterization of assimilatory nitrite reductase from Haloferax alexandrinus

It is aimed to purify assimilatory nitrite reductase (NiR) and to determine biochemical properties of this enzyme from halophilic archaeon Haloferax alexandrinus in this thesis. Assimilatory nitrite reductase from Haloferax alexandrinus was purified 34,5-fold. Sodium dodecyl sulfate – polyacrylamide gel electrophoresis (SDS-PAGE) analysis of the purified enzyme showed that NiR was a monomer with molecular weight of 62 kDa. Km values of H. alexandrinus for nitrite and methyl viologen were 8,25 mM and 1,46 mM respectively. The optimal pH value of the assimilatory nitrite reductase was 7.5 and the optimal temperature was 60°C. Maximum NiR activity was measured at 3.5 M NaCl concentration and the enzyme activity was doubled in the presence of KCl at the same concentrations. NiR enzyme activity of H. alexandrinus decreased in the presence of cyanide, azide, ethylenediamine tetra acetic acid, chlorate, and sulphite. However dithiothreitol had no effect on the activity of the enzyme.