Abstract
Catharsius protease-2 (CPM-2) was isolated from the body of dung beetles,Catharsius molossus, using a three step purification process (ammonium sulfate fractionation, gel filtration on Bio-Gel P-60, and affinity chromatography on DEAE Affi-Gel blue). The purified CPM-2, having a molecular weight of 24 kDa, was assessed homogeneously by SDS-polyacrylamide gel electrophoresis. TheN-terminal amino acid sequence of CPM-2 was composed of X Val Gln Asp Phe Val Glu Glu Ile Leu. CPM-2 was inactivated by Cu2+ and Zn2+ and strongly inhibited by typical serine proteinase inhibitors such as TLCK, soybean trypsin inhibitor, aprotinin, benzamidine, and α1-antitrypsin. However, EDTA, EGTA, cysteine, β-mercaptoethanol, E64, and elastatinal had little effect on enzyme activity. In addition, antiplasmin and antithrombin III were not sensitive to CPM-2. Based on the results of a fibrinolytic activity test, CPM-2 readily cleaved Aα-and Bβ-chains of fibrinogen and fibrin, and γ-chain of fibrinogen more slowly. The nonspecific action of the enzyme resulted in extensive hydrolysis, releasing a variety of fibrinopeptides of fibrinogen and fibrin. Polyclonal antibodies of CPM-2 were reactive to the native form of antigen. The ELISA was applied to detect quantities, in nanograms, of the antigen in CPM-2 protein.
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Ahn, M.Y., Hahn, BS., Ryu, K.S. et al. Purification and characterization of a serine protease (CPM-2) with fibrinolytic activity from the dung beetles. Arch Pharm Res 28, 816–822 (2005). https://doi.org/10.1007/BF02977348
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DOI: https://doi.org/10.1007/BF02977348