ISOLATION AND CHARACTERIZATION OF NITRILASE OF THERMOPHILIC PSEUDOMONAS SP.KNB-2, ISOLATED FROM INDUSTRIAL WASTE DISPOSAL SITE OF KIRTI NAGAR, NEW DELHI

Abstract

Microbial nitrilases are biocatalysts of industrial interest and being used for the production of commercial and pharmaceutically important products. In the present research work, an inducible thermophilic nitrilase containing strain was isolated by enrichment method from the dump site of Kirti Nagar, Delhi and was designated as Pseudomonas sp. KNB-2. The effect of different nitriles and amides, different medium composition, pH, temperature, inducer concentration and time course of incubation on production of nitrilase was optimized. It was found that benzonitrile (inducer) in the concentration of 0.3% (v/v), pH 7 and 55ᵒC in M1 medium were most suitable conditions for maximum synthesis of nitrilase and produced dry cell biomass of 16.24 g/L. Nitrilase exhibited temperature stability in wide range of 45 ᵒC to 65ᵒC and had optimum temperature of 45ᵒC, pH7 and exhibited maximum activity in benzonitrile, though it also catalyzed the conversion of acrylonitrile and acetonitrile. Nitrilase activity was observed in 30-40% cut of ammonium sulphate precipitation and two bands of molecular weight 43 kDa and 47 kDa were observed in SDS-PAGE analysis.