Purification and characterization of thermostable l-asparaginase from Bacillus amyloliquefaciens MKSE in Korean soybean paste

This study aimed to produce and characterize thermostable l-asparaginase from Bacillus amyloliquefaciens MKSE, a newly screened strain from Korean fermented soybean paste. The isolated MKSE was identified by its morphological, biochemical, and physiological properties and 16S rRNA sequence. Its l-asparaginase II-like gene was expressed in Escherichia coli BL21 (DE3), by using the pET-28a (+) vector. The produced l-asparaginase (specific activity of 4.75 IU/mg) was purified by a one-step procedure, using nickel-affinity chromatography, and its molecular weight of 38.2 kDa was determined by gel electrophoresis. Its optimum pH and temperature were 8.5 and 65 °C, respectively. The enzyme was highly stable in the pH range of 3.0–11.0 for 6 h incubation, and the enzyme retained about 57% of its activity at 95 °C for 6 h. Its activity was activated in the presence of Mn2+, Mg2+, and Fe2+ while it was inhibited by Zn2+, Cu2+, PMSF, and SDS. Its Km was 1.5 mM, and its Vmax was 136.3 IU/mg. The results of this study reveal that MKSE l-asparaginase has a great potential to be used in various industries, due to its stability at high temperature and over a wide pH range.