ABSTRACT
Azo dyes and nitro-aromatic compounds are the largest group of pollutants released in the environment as industrial wastes. They create serious health and environmental problems. Azoreductases catalyze the reduction of azo dyes and nitro compounds to their respective amines. AN azoreductase was purified up to 12-fold from Lysinibacillus sphaericus using ion-exchange and size exclusion chromatography. It was optimally active at pH 7.4 and 75 °C. It was stable at 70 °C for 30 min. The purified enzyme utilized NADH rather than NADPH as an electron donor to reduce substrates. The molecular weight of the purified enzyme was ~29 kDa. The enzyme also acted as nitroreductase and could selectively reduce the nitro group of 2-nitrophenol, 4-nitrobenzoic acid, 2-nitro-benzaldehyde and 3-nitrophenol. Reduction products of these compounds were identified by IR and NMR.
Subject(s)
Azo Compounds/metabolism , Bacillaceae/enzymology , NADH, NADPH Oxidoreductases/metabolism , Nitro Compounds/metabolism , Azo Compounds/analysis , Biotransformation , Hydrogen-Ion Concentration , NAD , NADH, NADPH Oxidoreductases/chemistry , NADH, NADPH Oxidoreductases/isolation & purification , NADP , Nitro Compounds/analysis , Nitroreductases , TemperatureABSTRACT
Azoreductase plays a key role in bioremediation and biotransformation of azo dyes. It initializes the reduction of azo bond in azo dye metabolism under aerobic or anaerobic conditions. In the present study, we isolated an alkaliphilic red-colored Aquiflexum sp. DL6 bacterial strain and identified by 16S rRNA method. We report nicotinamide adenine dinucleotide and nicotinamide adenine dinucleotide phosphate-dependent azoreductase purified from Aquiflexum sp. DL6 by a combination of ammonium sulfate precipitation and chromatography methods. The azoreductase was purified up to 30-fold with 37 % recovery. The molecular weight was found to be 80 kDa. The optimum activity was observed at pH 7.4 and temperature 60 °C with amaranth azo dye as a substrate. The thermal stability of azoreductase was up to 80 °C. The azoreductase has shown a wide range of substrate specificity, including azo dyes and nitro aromatic compounds. Metal ions have no significant inhibitory action on azoreductase activity. The apparent K m and V max values for amaranth azo dye were 1.11 mM and 30.77 U/mg protein respectively. This NAD (P) H azoreductase represents the first azoreductase to be characterized from alkaliphilic bacteria.
