Dramatic and concerted conformational changes enable rhodocetin to block α2β1 integrin selectively.

The collagen binding integrin alpha 2 beta 1 plays a crucial role in hemostasis, fibrosis, and cancer progression amongst others. It is specifically inhibited by rhodocetin (RC), a C-type lectin-related protein (CLRP) found in Malayan pit viper (Calloselasma rhodostoma) venom. The structure of RC alone reveals a heterotetramer arranged as an alpha beta and gamma delta subunit in a cruciform shape. RC specifically binds to the collagen binding A-domain of the integrin alpha 2 subunit, thereby blocking collagen-induced platelet aggregation. However, until now, the molecular basis for this interaction has remained unclear. Here, we present the molecular structure of the RC gamma delta-alpha 2A complex solved to 3.0 angstrom resolution. Our findings show that RC undergoes a dramatic structural reorganization upon binding to alpha 2 beta 1 integrin. Besides the release of the nonbinding RC alpha beta tandem, the RC gamma subunit interacts with loop 2 of the alpha 2A domain as result of a dramatic conformational change. The RC delta subunit contacts the integrin alpha 2A domain in the "closed" conformation through its helix C. Combined with epitope-mapped antibodies, conformationally locked a2A domain mutants, point mutations within the alpha 2A loop 2, and chemical modifications of the purified toxin protein, this molecular structure of RC gamma d-alpha 2A complex explains the inhibitory mechanism and specificity of RC for alpha 2 beta 1 integrin.