title: Proteolytic modulation of factor Xa-antithrombin complex enhances fibrinolysis in plasma.
abstract: Our previous work showed that purified coagulation factor Xa (FXa) acquires fibrinolysis cofactor activity after plasmin-mediated cleavage. The predominant functional species is a non-covalent heterodimer of 33 and 13kDa, termed Xa33/13, which has predicted newly exposed C-terminal lysines that are important for tissue plasminogen activator (tPA)-mediated plasminogen activation to plasmin. To provide evidence that this mechanism occurs in a physiological context, here we demonstrated the appearance of Xa33 in clotting plasma by western blot analysis. Since the normal fate of FXa is stable association with antithrombin (AT), an AT western blot was conducted, which revealed a band of ~13kDa higher apparent molecular weight than AT that appeared concurrent to Xa33. Sequencing of purified proteins confirmed the generation of Xa13 covalently bound to AT and Xa33 (Xa33/13-AT) by cleavages at Lys-Met339 and Lys-Asp389. Ligand blots demonstrated (125)I-plasminogen binding to the Xa33 subunit of plasmin-generated Xa33/13-AT. Purified XaAT added to plasma that was induced to clot enhanced the rate of tPA-mediated fibrinolysis by ~16-fold. Similarly, purified plasminogen activation by tPA was enhanced by ~16-fold by XaAT. Plasmin cleaves XaAT and exposes plasminogen binding sites at least 10-fold faster than FXa. Here we demonstrate a novel function for AT, which accelerates the modulation of FXa into the fibrinolytic form, Xa33/13. The consequent exposure of C-terminal lysine binding sites essential for plasminogen activation enhances fibrinolysis. These results are consistent with a model where auxiliary cofactors link coagulation to fibrinolysis by priming the accelerating role of fibrin.
authors: Department of Pathology and Laboratory Medicine, University of British Columbia, Canada.
authors: Kimberley
authors: K
authors: Talbot
authors: Scott C
authors: SC
authors: Meixner
authors: Edward L G
authors: EL
authors: Pryzdial
mesh_terms: D000991:Antithrombins / Q000097:blood*; D001665:Binding Sites; D001777:Blood Coagulation / Q000502:physiology; D015951:Factor Xa / Q000378:metabolism*; D005337:Fibrin / Q000378:metabolism; D005341:Fibrinolysin / Q000378:metabolism; D005342:Fibrinolysis / Q000502:physiology*; D006801:Humans; D010958:Plasminogen / Q000378:metabolism; D011485:Protein Binding; D059748:Proteolysis; D010959:Tissue Plasminogen Activator / Q000378:metabolism
chemical_list: D000991:Antithrombins; D005337:Fibrin; D010958:Plasminogen; D015951:Factor Xa; D010959:Tissue Plasminogen Activator; D005341:Fibrinolysin