Abstract
Covalent cross-linking of fibrin by the plasma transglutaminase coagulation factor XIII (FXIII) is a key determinant of blood clot stability and function. FXIII-catalyzed formation of ε-N-(γ-glutamyl)-lysyl cross-links is restricted to the fibrin γ-chain and α-chain and follows thrombin-driven fibrin polymerization. Fibrinogen is also cross-linked by tissue transglutaminase (TG2) in a reaction favoring intramolecular and intermolecular α-γ cross-linking. Emerging evidence indicates that fibrinogen is a relevant substrate of TG2 in conditions of acute tissue damage. Remarkably, beyond detection of prototypical FXIII-directed cross-links (ie, α-α, γ-γ), we identified entirely novel covalent cross-links involving the fibrinogen β-chain (ie, β-α, via FGB-Q82). Addition of TG2 to in vitro clotting reactions and analysis of fibrin(ogen) in reducing conditions revealed loss of β-chain polypeptide paired with formation of high-molecular weight β-chain species. Mass spectrometry-based cross-linking proteomic analysis of in vitro clots recapitulated the precise TG2-directed β-chain cross-links observed in clots made using plasma from patients following traumatic injury. The results indicate in vitro and ex vivo cross-linking of the fibrin β-chain and highlight a novel example of TG2 emerging as a relevant plasma transglutaminase.