Purification and characterization of a fibrinolytic enzyme from Streptomyces sp XZNUM 00004

A fibrinolytic enzyme (SFE1) from Streptomyces sp. XZNUM 00004 was purified to electrophoretic homogeneity with the methods including ammonium sulfate precipitation, polyacrylamide gel, DEAE-Sepharose Fast Flow anion exchange and gel-filtration chromatography. The molecular weight of SFE1 was estimated to be 20 kDa by SDS-PAGE, fibrin zymography, and gel filtration chromatography. The isoelectric point was 4.9. Km and Vmax values were 0.96 mg/ml and 181.8 unit/ml, respectively. It was very stable at pH 5.0-8.0 and below 65 degrees C. The optimum pH for enzyme activity was 7.8. The optimum temperature was 35 degrees C. The fibrinolytic activity of SFE1 was enhanced by Na+, K+, Mn2+, Mg2+, Zn2+ and Co2+. Conversely, Cu2+ showed strong inhibition. Furthermore, the fibrinolytic activity was strongly inhibited by PMSF, and partly inhibited by EDTA and EGTA. SFE1 rapidly hydrolyzed the Aa-chain of fibrinogen, followed by the Bb-chain and finally the c-chain. The first 15 amino acids of the N-terminal sequence were API-TLSQGHVDVVDI. Additionally, SFE1 directly digested fibrin and not by plasminogen activators in vitro. SFE1 can be further developed as a potential candidate for thrombolytic therapy.