Download PDF
Thromb Haemost 2001; 85(03): 458-463
DOI: 10.1055/s-0037-1615605
Review Article
Schattauer GmbH

The Role of Catalytic Cleft and Exosite Residues of Factor VIIa for Complex Formation with Tissue Factor Pathway Inhibitor

Alexei Iakhiaev
1   Department of Biochemistry, The University of Texas Health Center at Tyler, Tyler, Texas, and the Departments of Immunology and Vascular Biology, The Scripps Research Institute, La Jolla, California, USA
,
Wolfram Ruf
1   Department of Biochemistry, The University of Texas Health Center at Tyler, Tyler, Texas, and the Departments of Immunology and Vascular Biology, The Scripps Research Institute, La Jolla, California, USA
,
Vijaya Mohan L. Rao
1   Department of Biochemistry, The University of Texas Health Center at Tyler, Tyler, Texas, and the Departments of Immunology and Vascular Biology, The Scripps Research Institute, La Jolla, California, USA
› Author Affiliations
Further Information

Publication History

Received 21 September 2000

Accepted 19 October 2000

Publication Date:
08 December 2017 (online)

Also available at
    Permissions and Reprints

Summary

The extrinsic coagulation pathway is initiated by the binding of plasma factor VIIa (VIIa) to the cell surface receptor tissue factor (TF). Formation of the TF-VIIa complex results in allosteric activation of VIIa as well as the creation of an extended macromolecular substrate binding exosite that greatly enhances proteolytic activation of substrate factor X. The catalytic function of the TF-VIIa complex is regulated by a specific Kunitz-type inhibitor, tissue factor pathway inhibitor (TFPI). TFPI inhibition of the TF-VIIa complex was enhanced by the presence of Xa. This study investigates the relative contribution of catalytic cleft and exosite residues in VIIa for inhibitory complex formation with TFPI. VIIa protease domain residues Q176, T239 and E296 are involved in the formation of stable inhibitor complex with free TFPI. Kinetic analysis further demonstrated a predominant role of the S2’ subsite residue Q176 for the initial complex formation with TFPI. In contrast, no significant reductions in inhibition by TFPI-Xa were found for each of the mutants in complex with phospholipid reconstituted TF. However, reduced rates of inhibition of the VIIa Gla-domain (R36) and Q176 mutant by TFPI-Xa were evident when TF was solubilized by detergent micelles. These data demonstrate docking of the TFPI-Xa complex with the macromolecular substrate exosite and the catalytic cleft, in particular the S2’ subsite. The masking of the mutational effect by the presence of phospholipid shows a critical importance of Xa Gla-domain interactions in stabilizing the quaternary TF-VIIa-Xa-TFPI complex.

Keywords

Factor VIIa - tissue factor pathway inhibitor - tissue factor - factor Xa - protein interaction
 

  • References

  • 1 Rapaport SI, Rao LVM. The tissue factor pathway: How it has become a “prima ballerina”. Thromb Haemost 1995; 74: 7-17.
    PubMedGoogle Scholar
  • 2 Ruf W, Dickinson CD. Allosteric regulation of the cofactor dependent serine protease coagulation factor VIIa. Trends Cardiovasc Med 1998; 8: 350-6.
    CrossrefPubMedGoogle Scholar
  • 3 Betz A, Krishnaswamy S. Regions remote from the site of cleavage determine macromolecular substrate recognition by the prothrombinase complex. J Biol Chem 1998; 273: 10709-18.
    CrossrefPubMedGoogle Scholar
  • 4 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: 24171-5.
    CrossrefPubMedGoogle Scholar
  • 5 Dickinson CD, Kelly CR, Ruf W. Identification of surface residues mediating tissue factor binding and catalytic function of the serine protease. Proc Natl Acad Sci USA 1996; 93: 14379-84.
    CrossrefPubMedGoogle Scholar
  • 6 Ruf W, Miles DJ, Rehemtulla A, Edgington TS. Cofactor residues lysine 165 and 166 are critical for protein substrate recognition by the tissue factor-factor VIIa protease. J Biol Chem 1992; 267: 6375-81.
    PubMedGoogle Scholar
  • 7 Ruf W, Shobe J, Rao SM, Dickinson CD, Olson A, Edgington TS. Importance of factor VIIa Gla-domain residue Arg-36 for recognition of the macromolecular substrate factor X Gla-domain. Biochem 1999; 38: 1957-66.
    CrossrefPubMedGoogle Scholar
  • 8 Chang YZ, Hamaguchi N, Chang SC, Ruf W, Lin SW. Engineered recombinant factor VII Q217 variants with altered inhibitor specificities. Biochem 1999; 38: 10940-8.
    CrossrefPubMedGoogle Scholar
  • 9 Ruf W. Factor VIIa residue Arg290 is required for efficient activation of the macromolecular substrate factor X. Biochem 1994; 33: 11631-6.
    CrossrefPubMedGoogle Scholar
  • 10 Broze Jr GJ, Warren LA, Novotny WF, Higuchi DA, Girard JJ, Miletich JP. The lipoprotein-associated coagulation inhibitor that inhibits the factor VII-tissue factor complex also inhibits factor Xa: insight into its possible mechanism of action. Blood 1988; 71: 335-43.
    PubMedGoogle Scholar
  • 11 Rapaport SI. The extrinsic pathway inhibitor: a regulator of tissue factor-dependent blood coagulation. Thromb Haemost 1991; 66: 6-15.
    Article in Thieme ConnectPubMedGoogle Scholar
  • 12 Warn-Cramer BJ, Rao LVM, Maki SL, Rapaport SI. Modifications of extrinsic pathway inhibitor (EPI) and factor Xa that affect their ability to interact and to inhibit factor VIIa/tissue factor: Evidence for a two-step model of inhibition. Thromb Haemost 1988; 60: 453-6.
    Article in Thieme ConnectPubMedGoogle Scholar
  • 13 Broze Jr GJ, Girard TJ, Novotny WF. Regulation of coagulation by a multivalent Kunitz-type inhibitor. Biochem 1990; 29: 7539-45.
    CrossrefPubMedGoogle Scholar
  • 14 Girard TJ, Warren LA, Novotny WF, Likert KM, Brown SG, Miletich JP, Broze Jr GJ. Functional significance of the Kunitz-type inhibitory domains of lipoprotein-associated coagulation inhibitor. Nature 1989; 338: 518-20.
    CrossrefPubMedGoogle Scholar
  • 15 Girard TJ, MacPhail LA, Likert KM, Novotny WF, Miletich JP, Broze Jr GJ. Inhibition of factor VIIa-tissue factor coagulation activity by a hybrid protein. Science 1990; 248: 1421-24.
    CrossrefPubMedGoogle Scholar
  • 16 Callander NS, Rao LVM, Nordfang O, Sandset PM, Warn-Cramer B, Rapaport SI. Mechanisms of binding of recombinant extrinsic pathway inhibitor (rEPI) to cultured cell surfaces. Evidence that rEPI can bind to and inhibit factor VIIa-tissue factor complexes in the absence of factor Xa. J Biol Chem 1992; 267: 876-82.
    PubMedGoogle Scholar
  • 17 Hamamoto T, Yamamoto M, Nordfang O, Petersen JGL, Foster DC, Kisiel W. Inhibitory properties of full-length and truncated recombinant tissue factor pathway inhibitor (TFPI). Evidence that the third kunitz-type domain of TFPI is not essential for the inhibition of factor VIIa-tissue factor complexes on cell surfaces. J Biol Chem 1993; 268: 8704-10.
    PubMedGoogle Scholar
  • 18 Zhang E, St Charles R, Tulinsky A. Structure of extracellular tissue factor complexed with factor VIIa inhibited with a BPTI mutant. J Mol Biol 1999; 285: 2089-104.
    CrossrefPubMedGoogle Scholar
  • 19 Neuenschwander PF, Morrissey JH. Alteration of the substrate and inhibitor specificities of blood coagulation factor VIIa: importance of amino acid residue K192. Biochem 1995; 34: 8701-7.
    CrossrefPubMedGoogle Scholar
  • 20 Rao LVM, Ruf W. Tissue factor residues Lys165 and Lys166 are essential for rapid formation of quaternary complex of tissue factor. VIIa with Xa. tissue factor pathway inhibitor. Biochem 1995; 34: 10867-71.
    CrossrefPubMedGoogle Scholar
  • 21 Baugh RJ, Broze Jr GJ, Krishnaswamy S. Regulation of extrinsic pathway factor Xa formation by tissue factor pathway inhibitor. J Biol Chem 1998; 273: 4378-86.
    CrossrefPubMedGoogle Scholar
  • 22 Ruf W, Edgington TS. An anti-tissue factor monoclonal antibody which inhibits TF. VIIa complex is a potent anticoagulant in plasma. Thromb Haemost 1991; 66: 529-33.
    Article in Thieme ConnectPubMedGoogle Scholar
  • 23 Ruf W, Edgington TS. Two sites in the tissue factor extracellular domain mediate the recognition of the ligand factor VIIa. Proc Natl Acad Sci 1991; 88: 8430-4.
    CrossrefPubMedGoogle Scholar
  • 24 Stone MJ, Ruf W, Miles DJ, Edgington TS, Wright PE. Recombinant soluble human tissue factor secreted by Saccharomyces cerevisiae and refolded. Biochem J 1995; 310: 605-14.
    CrossrefPubMedGoogle Scholar
  • 25 Shobe J, Dickinson CD, Ruf W. Regulation of the catalytic function of coagulation factor VIIa by a conformational linkage of surface residue Glu 154 to the active site. Biochem 1999; 38: 2745-51.
    CrossrefPubMedGoogle Scholar
  • 26 Bajaj SP, Rapaport SI, Prodanos C. A simplified procedure for purification of human prothrombin, factor IX and factor X. Prep Biochem 1981; 11: 397-412.
    PubMedGoogle Scholar
  • 27 Rao LVM, Robinson T, Hoang AD. Factor VIIa/tissue factor-catalyzed activation of factors IX and X on a cell surface and in suspension: A kinetic study. Thromb Haemost 1992; 67: 654-9.
    Article in Thieme ConnectPubMedGoogle Scholar
  • 28 Lindhout T, Franssen J, Willems G. Kinetics of the inhibition of tissue factor-factor VIIa by tissue factor pathway inhibitor. Thromb Haemost 1995; 74: 910-15.
    Article in Thieme ConnectPubMedGoogle Scholar
  • 29 Jesty J, Wun TC, Lorenz A. Kinetics of the inhibition of factor Xa and the tissue factor-factor VIIa complex by the tissue factor pathway inhibitor in the presence and absence of heparin. Biochem 1994; 33: 12686-94.
    CrossrefPubMedGoogle Scholar
  • 30 Banner DW, D’Arcy A, Chene C, Winkler FM, Guha A, Konigsberg WH, Nemerson Y, Kirchhofer D. The crystal structure of the complex of blood coagulation factor VIIa with soluble tissue factor. Nature 1996; 380: 41-6.
    CrossrefPubMedGoogle Scholar
  • 31 Guex N, Peitsch MC. SWISS-MODEL and the Swiss-Pdb Viewer: an environment for comparative protein modeling. Electrophoresis 1997; 18: 2714-23.
    CrossrefPubMedGoogle Scholar
  • 32 Shah AM, Kisiel W, Foster DC, Nelsestuen GL. Manipulation of the membrane binding site of vitamin K-dependent proteins: enhanced biological function of human factor VII. Proc Natl Acad Sci 1998; 95: 4229-34.
    CrossrefPubMedGoogle Scholar
  • 33 Szedlacsek SE, Duggleby RG. Kinetics of slow and tight-binding inhibitors. Methods Enzymol 1995; 249: 144-80.
    PubMedGoogle Scholar
  • 34 Ruf W, Kalnik MW, Lund-Hansen T, Edgington TS. Characterization of factor VII association with tissue factor in solution. J Biol Chem 1991; 266: 15719-25.
    PubMedGoogle Scholar
  • 35 Lindhout T, Franssen J, Willems G. Kinetics of the inhibition of tissue factor-factor VIIa by tissue factor pathway inhibitor. Thromb Haemost 1995; 74: 910-5.
    Article in Thieme ConnectPubMedGoogle Scholar