Molecular, structural, and functional insights into the interaction of coagulation factor VIII with hemorrhage-derived heme

Introduction Hemarthrosis episodes in hemophilia A patients are associated with hemorrhagic events that lead to the breakdown of erythrocytes and, thus, the release and massive accumulation of labile heme. To avoid consequences, such as hemophilic arthropathy, exogenous substitution therapy with factor VIII (FVIII) is an option for affected patients. In this regard, it has been reported that direct FVIII injection into the joint bleeding, consequently into the heme-rich environment, shows significantly fewer side reactions. As such, FVIII and heme get in direct contact, however, basic information about this interaction is missing.

Method UV/Vis spectroscopic studies were used to characterize the heme-binding capacity of FVIII. To identify the respective heme-binding sites, FVIII was screened for potential heme-binding motifs by using the webserver HeMoQuest. Subsequently, these motifs were synthesized as FVIII-derived peptides and analyzed for heme binding using UV/vis spectroscopy. Promising sites were further evaluated by molecular docking simulations of the respective heme-protein complexes. Cryo-EM studies and clinical assay systems revealed functional aspects of heme binding to FVIII.

Results It is demonstrated that FVIII has with a binding capacity of seven heme molecules the so far highest known heme-binding capacity among all heme-binding proteins. The heme-binding regions are found in interaction sites. Furthermore, Cryo-EM studies as well as in silico studies suggest that heme binding induces higher flexibility in defined regions of FVIII. Finally, heme is thus able to diminish the procoagulant cofactor activity of FVIII.

Conclusion These results characterize FVIII as an extraordinary heme-binding protein and, thus, highlight the importance of the consideration of the FVIII-heme complex formation in the context of FVIII substitution therapy in hemophilia A patients with joint bleeding.

This research is funded by the Society for Thrombosis and Haemostasis Research e.V. (GTH) and the German Research Foundation (DFG).

Conflict of Interest

The authors declare no conflict of interest.

Publikationsverlauf

Artikel online veröffentlicht:
26. Februar 2024