A novel viper venom metalloproteinase, alborhagin, is an agonist at the platelet collagen receptor GPVI

The interaction of platelet membrane glycoprotein VI (GPVI) with collagen c an initiate (patho)physiological thrombus formation. The viper venom C-type lectin family proteins convulxin and alboaggregin-A activate platelets by interacting with GPVI. In this study, we isolated from white-lipped tree vi per (Trimeresurus albolabris) venom, alborhagin, which is functionally rela ted to convulxin because it activates platelets but is structurally differe nt and related to venom metalloproteinases. Alborhagin-induced platelet agg regation (EC50, <7.5 <mu>g/ml) was inhibitable by an anti-alpha IIb beta3 a ntibody, CRC64, and the Src family kinase inhibitor PP1, suggesting that al borhagin activates platelets, leading to alpha IIb,beta3-dependent aggregat ion. Additional evidence suggested that, like convulxin, alborhagin activat ed platelets by a mechanism involving GPVI. First, alborhagin- and convulxi n-treated platelets showed a similar tyrosine phosphorylation pattern, incl uding a similar level of phospholipase C gamma2 phosphorylation. Second, al borhagin induced GPVI-dependent responses in GPVI-transfected K562 and Jurk at cells. Third, alborhagin-dependent aggregation of mouse platelets was in hibited by the anti-GPVI monoclonal antibody JAQ1. Alborhagin had minimal e ffect on convulxin binding to GPVI-expressing cells, indicating that these venom proteins may recognize distinct binding sites. Characterization of al borhagin as a GPVI agonist that is structurally distinct from convulxin dem onstrates the versatility of snake venom toxins and provides a novel probe for GPVI-dependent platelet activation.