Cryo-EM structure of the tissue factor/factor VIIa complex with a factor X mimetic reveals a novel allosteric mechanism
- PMID: 40811856
- PMCID: PMC12397995
- DOI: 10.1182/blood.2025029430
Cryo-EM structure of the tissue factor/factor VIIa complex with a factor X mimetic reveals a novel allosteric mechanism
Abstract
Blood clotting is triggered in hemostasis and thrombosis when the membrane-bound tissue factor (TF)/factor VIIa (FVIIa) complex activates factor X (FX). There are no structures of TF/FVIIa on membranes, with or without FX. Using cryo-EM to address this gap, we assembled TF/FVIIa complexes on nanoscale membrane bilayers (nanodiscs), bound to XK1 and an antibody fragment. XK1 is a FX mimetic whose protease domain is replaced by the first Kunitz-type (K1) domain of tissue factor pathway inhibitor, while 10H10 is a non-inhibitory, anti-TF antibody. We determined a cryo-EM structure of a TF/FVIIa/XK1/10H10/nanodisc complex with a resolution of 3.7 Å, allowing us to model all the protein backbones. TF/FVIIa extends perpendicularly from the membrane, interacting with a "handle shaped" XK1 at two locations: the K1 domain docks into FVIIa's active site, while the γ-carboxyglutamate-rich (GLA) domain binds to TF's substrate-binding exosite. The FX and FVIIa GLA domains also contact each other and the membrane surface. Except for a minor contact between the first epidermal growth factor (EGF)-like domain of XK1 and TF, the rest of the FX light chain does not interact with TF/FVIIa. The structure reveals a previously unrecognized, membrane-dependent allosteric activation mechanism between FVIIa and TF where a serine-rich loop in TF that partially obscures the TF exosite must undergo a shift to allow access of the FX GLA domain to its final binding location on the membrane-bound TF/FVIIa complex. This mechanism also provides a novel explanation for the otherwise puzzling phenomenon of TF encryption/decryption on cell surfaces.
Copyright © 2025 American Society of Hematology.
Conflict of interest statement
Conflict-of-interest disclosure: All the authors declare no competing financial interests.
Figures





References
-
- Grover SP, Mackman N. Tissue factor: an essential mediator of hemostasis and trigger of thrombosis. Arteriosclerosis, Thrombosis and Vascular Biology. 2018;38(4):709–725. - PubMed
-
- Baugh RJ, Dickinson CD, Ruf W, Krishnaswamy S. Exosite interactions determine the affinity of factor X for the extrinsic Xase complex. J Biol Chem. 2000;275(37):28826–28833. - PubMed
-
- Shobe J, Dickinson CD, Edgington TS, Ruf W. Macromolecular substrate affinity for the tissue factor-factor VIIa complex is independent of scissile bond docking. J Biol Chem. 1999;274:24171–24175. - PubMed
-
- Roy S, Hass PE, Bourell JH, Henzel WJ, Vehar GA. Lysine residues 165 and 166 are essential for the cofactor function of tissue factor. J Biol Chem. 1991;266(32):22063–22066. - PubMed
Grants and funding
LinkOut - more resources
Full Text Sources
Miscellaneous