PURIFICATION FROM BOTHROPS LANCEOLATUS (FER DE LANCE) VENOM OF A FIBRINO(GENO)LYTIC ENZYME WITH ESTEROLYTIC ACTIVITY

Bothrops lanceolatus venom has high caseinolytic, phospholipasic, este rolytic and hemorrhagic activities. In spite of having no coagulant ef fect on plasma, this venom contains a thrombin-like enzyme. Using gel filtration and ion-exchange chromatographies, we have purified an este rolytic fraction (F-II-la) from this venom with a protein yield of 4% and a 58% recovery in enzyme activity. SDS-PAGE in the presence of bet a-mercaptoethanol showed that the enzyme is a single chain polypeptide with a M-w = 38,100. Immunodiffusion and immunoelectrophoresis of fra ction F-II-la against serum from horses immunized with B. lanceolatus venom and against rabbit antiserum prepared using fraction F-II-la bot h showed a single immunoprecipitin line. The K-m and V-max values for TAME hydrolysis were 0.85 mM and 38.6 mu mol/min/mg, respectively. The esterolytic activity was completely inhibited by PMSF (10 mM) but not by EDTA (20 mM). Fraction F-II-la hydrolyzed the alpha and beta chain s of fibrinogen. Degradation of the a chain occurred within 10 min whi le that of the beta-chain was slower. The enzyme had no effect on the gamma-chain even after 4 h of hydrolysis. (C) 1998 Elsevier Science Lt d. All rights reserved.