Mechanism of Factor Va Inactivation by Plasmin

The coagulation cofactor Va (FVa) is a noncovalent heterodimer consisting of a heavy chain (FVaH) and a light chain (FVaL). Previously, the fibrinolytic effector plasmin (Pn) has been shown to inhibit FVa function. To understand this mechanism, the fragmentation profile of human FVa by Pn and the noncovalent association of the derived fragments were determined in the presence of Ca2+ using anionic phospholipid (aPL)-coated microtiter wells and large (1 μm) aPL micelles as affinity matrices. Following Pn inactivation of aPL-bound FVa, a total of 16 fragments were observed and their NH2 termini sequenced. These had apparent molecular weights and starting residues as follows (single letter abbreviation is used): 50(L1766), 48(L1766), 43(Q1828), 40(Q1828), 30(S1546), 12(T1657), and 7(S1546) kDa from FVaL; and 65(A1), 50(A1), 45(A1), 34(S349), 30(L94), 30(M110), and 3 small <5(W457, W457, and K365) kDa from FVaH. Of these, 50(L1766), 48(1766), 43(Q1828), and 40(Q1828) spanning the C1/C2 domains, and 30(L94), but not the similar 30(M110), positioned within the A1 domain remained associated with aPL. These were detected antigenically during Pn- or tissue plasminogen activator-mediated lysis of fibrin clot formed in plasma. Chelation by EDTA dissociated the 30(L94)-kDa fragment, which was observed to associate with intact FVaL upon recalcification, indicating that the Leu-94 to Lys-109 region of the A1 domain plays a critical role in the FVaL and FVaH Ca2+-dependent association. By using domain-specific monoclonal antibodies and an assay for thrombin generation, loss of FVa prothrombinase function was coincident with proteolysis at sites in the A2 and A3 domains resulting in their dissociation. Inactivation of FV or FVa by Pn was independent of the thrombophilic R506Q mutation. These results identify the molecular composition of Pn-cleaved FVa that remains bound to membrane as largely A1-C1/C2 in the presence of Ca2+ and suggest that Pn inhibits FVa by a process involving A2 and A3 domain dissociation.

• Publication year

  2001

• Venue

  Journal of Biological Chemistry

• Publication date

  2001-06-08

• Fields of study

  Medicine, Chemistry

• Identifiers
  DOI 10.1074/jbc.M004711200PMID 11278280
• External record

  Open on Semantic Scholar

• Source metadata

  Semantic Scholar, PubMed

• No claims are published for this paper.

• No concepts are published for this paper.

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