A novel type of protease (mantis egg fibrinolytic enzyme, MEF-2) was isolated from the egg cases of Tenodera sinensis. The protease was homogeneous by SDS–PAGE and its apparent molecular mass was 32,900 Da. The amino acids in the N-terminal region were Ile–Val–Gly–Gly–Glu–Glu–Ala–Val–Ala–Gly–Asp–Phe–Pro–Ile–Val–Ser–Leu–Gln–Glu. The enzyme was inhibited by PMSF, TLCK, aprotinin, benzamidine, soybean trypsin inhibitor and also slightly by elastatinal, EDTA, EGTA, cysteine and β-mercaptoethanol, but TPCK, iodoacetate and E-64 did not affect the activity. MEF-2 was not sensitive to α 1-antitrypsin but antithrombin III and α 2-antiplasmin inhibited the enzyme. MEF-2 preferentially cleaved the oxidized B-chain of insulin between Arg 22 and Gly 23. Among chromogenic protease substrates, the most susceptible to MEF-2 hydrolysis was benzoyl-Phe–Val–Arg- p-nitroanilide with maximal activity at 30°C and pH 5.0. These results indicate that MEF-2 belongs to the trypsin family. Upon incubation of crosslinked fibrin with MEF-2, a steady increase of D-dimer suggests that the enzyme has a strong fibrinolytic activity. In conclusion, MEF-2 is a new type of proteolytic enzyme and has some potential for practical application in fibrinolysis.