Biochemical characterization of lebetase, a direct-acting fibrinolytic enzyme from Vipera lebetina snake venom.

Lebetase, the fibrinolytic enzyme isolated from Vipera lebetina (Levantine viper) snake venom is a metalloenzyme that contains one mole of Zn2+ and one mole of Ca2+ per mole of protein. Lebetase is inhibited by dithiothreitol, suggesting that disulfide bonds are necessary for holding the structure. Vipera lebetina venom contains several isoforms of lebetase in the interval of pI 4.6-5.4. Two lebetase fractions I (pI of the main component 5.0) and II (pI of the main component 5.3) degrade fibrin and fibrinogen by hydrolysis of the alpha and beta chains. The molecular weights of the cleavage products produced by the two different lebetase fractions are identical. The metal ions, Cd2+, Cu2+, Co2+, inhibit fibrinolytic and caseinolytic activity of lebetase I and II. Using mass spectrometry we characterized differences in molecular masses of lebetase I and II (22719 Da and 22912 Da). Vipera lebetina venom from a single snake contains mainly one form of lebetase. Lebetase I is more stable at low pH than lebetase II. The lebetases I and II inhibit platelet aggregation induced by ADP in a dose-dependent manner.