Production Purification and Characterization of Extracellular Anti-Leukaemic L-Asparaginase from Isolated Bacillus Subtilis Using Solid State Fermentation.

ABSTRACT

Bacterial L-asparaginase has been widely used as therapeutic agent in treatment of various lymphoblastic leukemia and food processing aid to reduce the formation of cancer causing acrylamide. The present work deals with production and purification of extracellular L-asparaginase from soil isolates using solid state fermentation. The isolate was characterized by big chemical test and identified as Bacillus subtilis. The enzyme production was carried out by solid state fermentation comparing the results with submerged fermentation. The enzyme was purified to near homogeneity by ammonium sulphate precipitation, dialysis, followed by gel filtration on Sephadex G-100 column, CM Sephadex C-50 and SDS-PAGE. The enzyme was purified at 110.2 folds and showed a final specific activity of 1785.7 IU/mg with 26.5% yield. SDS-PAGE of the purified enzyme revealed an apparent molecular weight of 109 kDa. The purified enzyme showed maximum activity at pH 9 when it was incubated at 50°C for 35 min. The enzyme was activated by Mg+2 and strongly inhibited by EDTA.