A hemorrhagin as a metalloprotease in the venom of Trimeresurus purpureomaculatus: purification and characterization.

A major hemorrhagin was purified from the venom of the Thai green pit viper (Trimeresurus purpureomaculatus) by gel filtration, ion-exchange and affinity chromatography. A 15-fold purification was achieved with an overall yield of 7% of hemorrhagic activity. The hemorrhagin was homogeneous according to disc- and SDS-PAGE as well as on immunodiffusion. The molecular weight determined by SDS-PAGE was 72kDa. The purified hemorrhagin expresses proteolytic activity with heat-denatured casein and hide powder azure, but it was free of AE-hydrolase and phospholipase activities. Both hemorrhagic and proteolytic activities were inhibited by EDTA, suggesting that the hemorrhagin is a metalloprotease. The hemorrhagin hydrolyzed all gelatin preparations derived from types I, II, III and IV collagen, whereas it hydrolyzed only type IV native collagen. The hemorrhagic activity was neutralized by Thai green pit viper antivenom raised to Trimeresurus albolabris venom.