Purification and properties of a thermostable phytase from Bacillus sp. DS11

Abstract

Bacillus species producing a thermostable phytase was isolated from the soil of a Korean cattle shed. An extracellular phytase from Bacillus sp. DS11 was purified to homogeneity by acetone precipitation and Phenyl-Sepharose, Resource S, and Superose 12 column chromatographies. Its molecular weight was estimated to be 44 kDa by SDS-polyacrylamide gel electrophoresis. Its optimum temperature for phytase activity was 70°C and about 50% of its original activity remained after incubation at 90°C for 1O min in the presence of 5 mM CaCl2. Calcium ions were required for thermal stability. The optimum pH for enzyme activity was 7.0 and fairly stable from pH 4.0-8.0. The enzyme had an isoelectric point of 5.3. The K(m) value for phytate was 0.55 mM. Its activity was greatly inhibited by EDTA and metal ions such as Cd2+ and Mn2+. As for substrate specificity it was very specific for phytate and had little or no activity on other phosphate esters. The enzyme efficiently hydrolyzed phytate in rice flour.