2 VASCULAR APOPTOSIS-INDUCING PROTEINS FROM SNAKE-VENOM ARE MEMBERS OF THE METALLOPROTEASE DISINTEGRIN FAMILY/

Hemorrhagic snake venom induces apoptosis in vascular endothelial cell s. In a previous report, we described the purification of a vascular a poptosis-inducing protein (VAP) from Crotalus atrox [Masuda, S., Araki , S., Kaji, K. & Hayashi, H. (1997) Biochem. Biophys. Res, Commun. 235 , 59-63]. We report here the identification of a second vascular apopt osis-inducing protein, VAP2, in venom from C. atrox. When we fractiona ted crude venom from C. atrox by isoelectric focusing, we found two pr oteins with apoptosis-inducing activity, one was basic and the other a cidic. The basic protein corresponded to VAP, and we named the acidic protein VAP2. VAP2 was a monomeric protein with molecular mass of 55 k Da and an isoelectric point of 4.5. VAP2 killed vascular endothelial c ells in culture, and the death of cells exhibited the characteristic f eatures of apoptotic activity of VAP2, seemed to be specific to endoth elial cells, as reported for VAP. The half-lethal doses of VAP and VAP 2 were 0.3 mu g/ml and 0.1 mu g/ml, respectively. Analysis of the part ial amino acid sequences of VAP and VAP2 revealed similarities to memb ers of the metalloprotease/disintegrin family. The sensitivity of VAP2 to changes in pH and temperature was distinct from that of VAP. Our r esults suggest that VAP and VAP2 are members of the metalloprotease/di sintegrin family.