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 alpha-chain followed by beta-chain. PTEFP rapidly degraded A alpha-chain of human fibrinogen but did not hydrolyze B beta- or gamma-chain indicating that it is alpha-fibrinogenase. The N-terminal sequence was AQNIGAVVNLSPPKQ which is different from that of other known fibrinolytic enzymes. The PTEFP displayed maximum activity at 35 degrees C and pH 5.0, and was stable between pH 5.0-8.0 and below 40 degrees 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 K-m and V-max 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. (C) 2011 Elsevier Ltd. All rights reserved.