Biochemical characteristics of a fibrinolytic enzyme purified from a marine bacterium, Bacillus subtilis HQS-3

Abstract A fibrinolytic enzyme isolated from marine Bacillus subtilis HQS-3 was purified to electrophoretic homogeneity using ammonium sulphate precipitation, alkaline solution treatment, membrane concentration, dialysis, ion exchange, and gel filtration chromatography. SDS-PAGE and gel filtration chromatography showed that it was a monomeric protein with an apparent molecular weight of 26 kDa. The purified enzyme was active at pH 6.0–10.0 with an optimum pH of 8.0. It was stable at temperatures ranging from 25 to 37 °C, exhibiting maximum activity between 45 °C and 50 °C. The isoelectric point of the enzyme was 9.0–9.2, which was higher than those of other known fibrinolytic enzymes from Bacillus species. PMSF, EDTA, Cu 2+ , Zn 2+ , and Co 2+ inhibited the enzyme activity significantly. This enzyme did not cause hemolysis in vitro and preferred direct degradation of fibrin in the following order: α, β, and γ–γ chains. Thus, these results suggest that the marine-derived enzyme is a plasmin-like serine metalloprotease, which is distinct from other fibrinolytic enzymes from genus Bacillus .