Intracellular L-Asparaginase from Bacillus sp. PG02: Purification, Biochemical Characterization and Evaluation of Optimum pH and Temperature

Abstract

Bacterial L-asparaginases are amidohydrolases that act on L-asparagine and produce L-aspartate and ammonia. These enzymes have been used in treatment of lymphoblastic leukemia. In the present study, a novel strain, Bacillus sp. PG02 was explored for the production of intra-cellular L-asparaginase enzyme. The nitrogen source for L-asparaginase production was L-asparagine. New intracellular L-asparaginase was purified using ion exchange chromatography and the purity was assessed using SDS-PAGE. Kinetic parameters k_m and V_max and thermal properties were studied using L-asparagine as the substrate. SDS-PAGE analysis showed apparent molecular weight of approximately 38 kDa. The enzyme was active in a wide pH ranges (5-10) and it was maximally active at pH 7.5. Bacillus PG02 L-asparaginase was optimally active at 40°C. Thermal inactivation studies exhibited t_1/2 of 32.5 min in 37°C. Also T50 and △G of inactivation were measured. The results revealed that the enzyme had appropriate characteristics and thus could be a potential candidate for medical and basic investigations.