Structural Superimposition

Abstact

BACKGROUND: Factor (F)XI occupies a unique and clinically significant niche, bridging the tissue factor-driven and contact-driven coagulation pathways. Irrespective of the trigger, activated factor XI (FXIa) contributes to clotting by activating FIX. Biochemical studies established that this reaction requires the membrane-binding FIX-Gla domain to engage an exosite on the FXIa Apple 3 (A3) domain, which only becomes available upon FXI activation. Structural data for FXIa, FIX, FIXabeta and the FXIa:FIX complex are lacking; current understanding relies on zymogen FXI crystal structures and homology modeling of FXIa after kallikrein. OBJECTIVES: Elucidate the high-resolution structure of FXIa in a functionally relevant conformation in complex with FIX. METHODS: We utilized cryo-electron microscopy to determine the structures of human FXIa in complex with its full-length substrate, FIX, and activated product, FIXabeta. RESULTS: We report the first cryo-electron microscopy structures of FXIa in complex with its substrate, FIX, and activated FIX (FIXabeta). The structures capture a functionally relevant conformational change in the FXIa catalytic domain and reveal the first view of the entire FIX and FIXabeta. Critically, we visualize the FIX-Gla domain precisely docked to the FXIa-A3 exosite on both subunits of the FXIa dimer. We also define the first step of proteolysis, visualizing the FIX Arg145 inserted into the primary specificity pocket of FXIa. CONCLUSIONS: The structures define the full FXIa:FIX interface, providing a structural template for understanding the sequential activation of FIX and for developing a new class of selective allosteric antithrombotic agents.