Purification and characterization of a novel, highly potent fibrinolytic enzyme from Paecilomyces tenuipes

A fibrinolytic enzyme (PTEFP) was purified from the entomopathogenic fungus Paecilomyces tenuipes. Analysis of the purified PTEFP by SDS–PAGE and fibrin zymography demonstrated a single protein band of approximately 14 kDa. Fibrinolysis pattern showed that PTEFP rapidly hydrolyzed α-chain followed by β-chain. PTEFP rapidly degraded Aα-chain of human fibrinogen but did not hydrolyze Bβ- or γ-chain indicating that it is α-fibrinogenase. The N-terminal sequence was AQNIGAVVNLSPPKQ, which is different from that of other known fibrinolytic enzymes. The PTEFP displayed maximum activity at 35 °C and pH 5.0, and was stable between pH 5.0–8.0 and below 40 °C. Calcium ion enhanced the enzyme activity whereas Zn2+ inhibited it. The fibrinolytic activity was strongly inhibited by PMSF identifying it as a serine protease. PTEFP exhibited high specificity for the substrate H-D-Val-Leu-Lys-pNA and Km and Vmax values for this substrate were 0.17 mM and 59 U/ml respectively. These results suggest that PTEFP is a novel fibrinolytic enzyme and may have potential applications in treating thrombosis.