CROTAVIRIN, A POTENT PLATELET-AGGREGATION INHIBITOR PURIFIED FROM THEVENOM OF THE SNAKE CROTALUS-VIRIDIS

A potent platelet aggregation inhibitor in the venom of Crotalus virid is snake was purified to homogeneity by gel filtration chromatography and reverse phase high-performance liquid chromatography. This purifie d principle, named crotavirin, is a single-chain polypeptide with a me l. wt of 9200 as estimated by SDS-polyacrylamide gel electrophoresis. It inhibited the aggregation of human washed platelets induced by coll agen, thrombin and thomboxane analogue (U46619) with a similar IC50 (s imilar to 1.0 mu g/ml, 0.11 mu M). The binding of fluorescein isothioc yanate-conjugated crotavirin to platelets was abolished in the presenc e of divalent cation chelator, EDTA, indicating that divalent cation i s essential for crotavirin's binding. A monoclonal antibody, 7E3, rais ed against platelet glycoprotein IIb-IIIa complex blocked the binding of fluorescein isothiocyanate-conjugated crotavirin to platelets, wher eas the other monoclonal antibody against glycoprotein IIb-IIIa, 10E5, had no inhibitory effect. In addition, crotavirin inhibited in a conc entration-dependent manner the binding of fluorescein isothiocyanate-c onjugated rhodostomin, a member of the disintegrin family, to platelet s. Its binding to platelets was blocked by disintegrins, e.g. trigrami n and rhodostomin. It is concluded that crotavirin is a potent platele t aggregation inhibitor, which acts specifically on an epitope of glyc oprotein IIb-IIIa, leading to the blockade of fibrinogen binding to gl ycoprotein IIb-IIIa and eventually the blockade of platelet aggregatio n.