Purification and characterization of a fibrinolytic enzyme produced from Enterococcus faecalis BRGA-5

Enterococcus faecalis BRCA-5, which produces a strong fibrinolytic enzyme, was screened from Tempe, a traditional Indonesian fermentedsoybean food. The enzyme was purified to homogeneity from culture broth of E. faecalis BRCA-5 by means of Chromatographie procedures and showed a strong fibrinolytic activity. The purified enzyme, named tempase, has a molecular weight of 38 kDa as determined by SDS-polyacrylamide gel electrophoresis under both reduced and non-reduced conditions, indicating that it consists of a single polypeptide chain. The enzyme displayed a very broad optimal pH range and its pi value was 4.8. Tempase was easily inhibited by metal-chelating agents but not by serine protease inhibitors, suggesting the enzyme belongs to metalloenzyme family. Furthermore, the inhibition of enzyme activity by chelating agents was recovered by the addition of divalent ions such as Ca2, Mg2, Co2 and Mn ⁺². The fibrinolytic activity was about 35 CU (plasmin units)/mg of protein as calculated from fibrin plate assay method.