L-Asparaginase from an acrylamide degrader, Cupriavidus oxalaticus ICTDB921: Production, kinetic modelling, purification and characterization

Abstract

In the present study, a novel soil isolate and previously reported acrylamide degrader, Cupriavidus oxalaticus ICTDB921, was found to have better and significantly higher l-asparaginase (LAase) activity than other bacterial strains that were screened for the same. The medium and fermentation conditions were optimized for maximizing LAase production followed by its purification and biochemical characterization. The optimum conditions of pH (7), temperature (30 °C) and agitation (180 rpm), showed the highest biomass accumulation (DCW 3.15 g/L) and LAase activity (22.94 IU/mL) after 12 h at 160 mM l-asparagine. Kinetic modelling of optimized batch data showed the biomass accumulation, LAase formation and l-asparagine utilization to best fit with the logistic model, Leudeking-Piret model and exponential decay model, respectively. The crude LAase from this isolate showed high potency at pH 7 to 9, and temperatures from 30 to 40 °C with reasonable pH (6–9) and thermal (up to 50 °C) stability. The LAase was stable against most amino acids and metal ions, and it could degrade aliphatic amides like urea and formamide. A sequential purification by ammonium sulphate precipitation, thermal treatment and chromatographic separation achieved approximately 8-fold purity. The purified subunit showed a molecular weight of 78 kDa, and km and Vmax of 24.1 mM and 40 μmol/min, respectively. The study demonstrates the potential of LAase from Cupriavidus oxalaticus in the mitigation of asparagine in food as well as for therapeutic applications.