Structural basis for pure antagonism of integrin αVβ3 by a high-affinity form of fibronectin

Integrins are important therapeutic targets. However, current RGD-based anti-integrin drugs are also partial agonists, inducing conformational changes that trigger potentially fatal immune reactions and paradoxical cell adhesion. Here we describe the first crystal structure of alpha(V)beta(3) bound to a physiologic ligand, the tenth type III RGD domain of wild-type fibronectin (wtFN10), or to a high-affinity mutant (hFN10) shown here to act as a pure antagonist. Comparison of these structures revealed a central pi-pi interaction between Trp1496 in the RGD-containing loop of hFN10 and Tyr122 of the beta(3) subunit that blocked conformational changes triggered by wtFN10 and trapped hFN10-bound alpha(V)beta(3) in an inactive conformation. Removing the Trp1496 or Tyr122 side chains or reorienting Trp1496 away from Tyr122 converted hFN10 into a partial agonist. These findings offer new insights into the mechanism of integrin activation and a basis for the design of RGD-based pure antagonists.