Abstract

Echistatin is a 49-amino-acid peptide belonging to the family of disintegrins that are derived from snake venoms and are potent inhibitors of platelet aggregation and cell adhesion. Integrin α_vβ₃ receptor plays a critical role in several physiological processes such as tumor-induced angiogenesis, tumor cell metastasis, osteoporosis and wound repair. In this study, we have characterized the binding of echistatin to purified integrin α_vβ₃ receptor and the form expressed on human embryonic kidney 293 cells. We show that both purified and membrane-bound integrin α_vβ₃ binds to echistatin with a high affinity, which can be competed efficiently by linear and cyclic peptides containing the RGD sequence. Previous studies have shown that α_vβ₃ binds to vitronectin in a nondissociable manner, whereas an RGD-containing peptide derived from vitronectin binds in a dissociable manner with aK_d of 9.4 × 10⁻⁷ M. Our studies indicate that radiolabeled echistatin binds to α_vβ₃ in a nondissociable manner, similar to native echistatin. However, echistatin does not support the adhesion of 293 cells expressing α_vβ₃ receptor because of poor binding to plastic dishes and is a potent antagonist of the adhesion of these cells to vitronectin. These studies demonstrate that echistatin binding to α_vβ₃ is of high affinity and irreversible similar to vitronectin and provides an alternate ligand for high-throughput screening for α_vβ₃ antagonists.