Functional Domains of Coagulation Factor V

Summary

Principal Investigator: M Kalafatis
Abstract: Prothrombinase is composed of the protein cofactor, factor Va, and the enzyme, factor Xa, associated on a cell surface in the presence of divalent metal ions. Incorporation of factor Va into prothrombinase and its interaction with factor Xa results in a 300,000-fold acceleration of the catalytic efficiency of the enzyme as compared to the catalysis of the reaction by factor Xa alone. The procofactor, factor V, does not participate in prothrombinase. Following activation of factor V by thrombin, factor Va is composed of heavy and light chains associated via divalent metal ions. Both chains of the cofactor interact with factor Xa while only the heavy chain of the cofactor binds prothrombin. The factor Va cofactor activity is efficiently down-regulated following proteolysis of the heavy chain by activated protein C (APC) only in the presence of a membrane surface and results in the inability of the cofactor to bind factor Xa. Thus, the positive and negative regulatory processes associated with factor V activation and its inactivation are directly associated with the capability of the cofactor to be incorporated into prothrombinase and to bind factor Xa. The amino acids responsible for the interaction of factor Va with factor Xa and prothrombin remain to be identified. We have data demonstrating that the heavy chain of the cofactor possesses a binding region for factor Xa within amino acid region 323-331, whereas the NH2-terminal portion of the light chain (amino acid residues 1546-1558) also interacts with factor Xa. In addition, we have data suggesting that the COOH-terminal portion of the heavy chain contain an interactive site for prothrombin while previous data have suggested that a binding site for thrombin is located on the B region of the procofactor. The specific aims of this grant proposal are: (1) to identify and characterize the factor Xa-binding domain(s) on factor Va light chain; (2) to identify and characterize the thrombin and prothrombin-binding domain(s) on the factor V molecule; (3) to test the physiological relevance of our findings by studying the assembly and function of prothrombinase on platelets. To achieve these goals we have designed a series of experiments that are prioritized and integrated with complementary molecular and structural approaches. Characterization of the specific amino acid regions of factor V that are critical for its function will allow for a profound understanding of the macromolecular interactions that control prothrombinase and are required for its assembly, function, and specificity. We have established a system to study phospholipids-driven macromolecular complex formation, which may be a model for the generation of complexes that form extra-and intra-cellularly.
Funding Period: 2004-05-01 - 2010-04-30
more information: NIH RePORT

Top Publications

  1. pmc Amino acid region 1000-1008 of factor V is a dynamic regulator for the emergence of procoagulant activity
    Joesph R Wiencek
    From the Department of Chemistry, Cleveland State University, Cleveland, Ohio 44115
    J Biol Chem 288:37026-38. 2013
  2. pmc Cleavage at both Arg306 and Arg506 is required and sufficient for timely and efficient inactivation of factor Va by activated protein C
    Melissa A Barhoover
    Department of Chemistry, Cleveland State University, 2351 Euclid Avenue, Cleveland, OH 44115, USA
    Blood Coagul Fibrinolysis 22:317-24. 2011
  3. pmc Contribution of amino acid region 659-663 of Factor Va heavy chain to the activity of factor Xa within prothrombinase
    JAMILA HIRBAWI
    Department of Chemistry, Cleveland State University, Cleveland, Ohio 44115, USA
    Biochemistry 49:8520-34. 2010
  4. pmc Cooperative regulation of the activity of factor Xa within prothrombinase by discrete amino acid regions from factor Va heavy chain
    Melissa A Barhoover
    Department of Chemistry, Cleveland State University, Cleveland, Ohio 44115, USA
    Biochemistry 47:12835-43. 2008
  5. pmc Role of the acidic hirudin-like COOH-terminal amino acid region of factor Va heavy chain in the enhanced function of prothrombinase
    JAMILA HIRBAWI
    Department of Chemistry, Cleveland State University, Cleveland, Ohio 44115, USA
    Biochemistry 47:7963-74. 2008
  6. pmc Contribution of amino acid region 334-335 from factor Va heavy chain to the catalytic efficiency of prothrombinase
    Melissa A Barhoover
    Department of Chemistry, Cleveland State University, Cleveland, Ohio 44115, USA
    Biochemistry 47:6840-50. 2008
  7. ncbi The interaction of fragment 1 of prothrombin with the membrane surface is a prerequisite for optimum expression of factor Va cofactor activity within prothrombinase
    Michael A Bukys
    Cleveland State University, Department of Chemistry, OH 44115, USA
    Thromb Haemost 99:511-22. 2008
  8. ncbi Identification of an inactivating cleavage site for alpha-thrombin on the heavy chain of factor Va
    Evrim Erdogan
    Department of Chemistry, Cleveland State University, 2351 Euclid Avenue, Science and Research Center SR 370, Cleveland, Ohio 44115, USA
    Thromb Haemost 98:998-1006. 2007
  9. ncbi The contribution of amino acid residues 1508-1515 of factor V to light chain generation
    E Erdogan
    Department of Chemistry, Cleveland State University, Cleveland, OH 44115, USA
    J Thromb Haemost 6:118-24. 2008
  10. ncbi A control switch for prothrombinase: characterization of a hirudin-like pentapeptide from the COOH terminus of factor Va heavy chain that regulates the rate and pathway for prothrombin activation
    Michael A Bukys
    Department of Chemistry, Cleveland State University, Cleveland, Ohio 44115, USA
    J Biol Chem 281:39194-204. 2006

Scientific Experts

  • M Kalafatis
  • Michael A Bukys
  • Melissa A Barhoover
  • Tivadar Orban
  • JAMILA HIRBAWI
  • Michael E Nesheim
  • Paul Y Kim
  • Evrim Erdogan
  • Joesph R Wiencek
  • E Erdogan
  • Daniel O Beck
  • Mahesheema Na
  • John L Vaughn
  • Hans L Vos
  • M A Bukys
  • Thomas Orfeo
  • Kenneth G Mann
  • Melissa A Blum
  • Nicole Brufatto
  • Valentin Gogonea

Detail Information

Publications14

  1. pmc Amino acid region 1000-1008 of factor V is a dynamic regulator for the emergence of procoagulant activity
    Joesph R Wiencek
    From the Department of Chemistry, Cleveland State University, Cleveland, Ohio 44115
    J Biol Chem 288:37026-38. 2013
    ....
  2. pmc Cleavage at both Arg306 and Arg506 is required and sufficient for timely and efficient inactivation of factor Va by activated protein C
    Melissa A Barhoover
    Department of Chemistry, Cleveland State University, 2351 Euclid Avenue, Cleveland, OH 44115, USA
    Blood Coagul Fibrinolysis 22:317-24. 2011
    ....
  3. pmc Contribution of amino acid region 659-663 of Factor Va heavy chain to the activity of factor Xa within prothrombinase
    JAMILA HIRBAWI
    Department of Chemistry, Cleveland State University, Cleveland, Ohio 44115, USA
    Biochemistry 49:8520-34. 2010
    ....
  4. pmc Cooperative regulation of the activity of factor Xa within prothrombinase by discrete amino acid regions from factor Va heavy chain
    Melissa A Barhoover
    Department of Chemistry, Cleveland State University, Cleveland, Ohio 44115, USA
    Biochemistry 47:12835-43. 2008
    ....
  5. pmc Role of the acidic hirudin-like COOH-terminal amino acid region of factor Va heavy chain in the enhanced function of prothrombinase
    JAMILA HIRBAWI
    Department of Chemistry, Cleveland State University, Cleveland, Ohio 44115, USA
    Biochemistry 47:7963-74. 2008
    ....
  6. pmc Contribution of amino acid region 334-335 from factor Va heavy chain to the catalytic efficiency of prothrombinase
    Melissa A Barhoover
    Department of Chemistry, Cleveland State University, Cleveland, Ohio 44115, USA
    Biochemistry 47:6840-50. 2008
    ..The data demonstrate that amino acid region 334-335 is required for the rearrangement of enzyme and substrate necessary for efficient catalysis of prothrombin by prothrombinase...
  7. ncbi The interaction of fragment 1 of prothrombin with the membrane surface is a prerequisite for optimum expression of factor Va cofactor activity within prothrombinase
    Michael A Bukys
    Cleveland State University, Department of Chemistry, OH 44115, USA
    Thromb Haemost 99:511-22. 2008
    ..Altogether, the data demonstrate that membrane-bound fragment 1 is required to promote optimum Fva cofactor activity which in turn is translated by efficient initial cleavage of prothrombin by prothrombinase at Arg(320)...
  8. ncbi Identification of an inactivating cleavage site for alpha-thrombin on the heavy chain of factor Va
    Evrim Erdogan
    Department of Chemistry, Cleveland State University, 2351 Euclid Avenue, Science and Research Center SR 370, Cleveland, Ohio 44115, USA
    Thromb Haemost 98:998-1006. 2007
    ..Our data demonstrate that cleavage of FVa at Arg(643) by alpha-thrombin results in a partially inactive cofactor molecule and provides for an activated protein C (APC)-independent anticoagulant effect of alpha-thrombin...
  9. ncbi The contribution of amino acid residues 1508-1515 of factor V to light chain generation
    E Erdogan
    Department of Chemistry, Cleveland State University, Cleveland, OH 44115, USA
    J Thromb Haemost 6:118-24. 2008
    ..Factor (F) V is activated by alpha-thrombin following cleavages at Arg(709), Arg(1,018) and Arg(1,545). Amino acid region 1,490-1,520 of FV is essential for procofactor activation...
  10. ncbi A control switch for prothrombinase: characterization of a hirudin-like pentapeptide from the COOH terminus of factor Va heavy chain that regulates the rate and pathway for prothrombin activation
    Michael A Bukys
    Department of Chemistry, Cleveland State University, Cleveland, Ohio 44115, USA
    J Biol Chem 281:39194-204. 2006
    ..Our data demonstrate that pentapeptide DYDYQ has opposing effects on membrane-bound factor Xa for prothrombin cleavage, depending on the incorporation of factor Va in prothrombinase...
  11. ncbi The structural integrity of anion binding exosite I of thrombin is required and sufficient for timely cleavage and activation of factor V and factor VIII
    Michael A Bukys
    Department of Chemistry, Cleveland State University, Cleveland, Ohio 44115, USA
    J Biol Chem 281:18569-80. 2006
    ....
  12. ncbi Completed three-dimensional model of human coagulation factor va. Molecular dynamics simulations and structural analyses
    Tivadar Orban
    Department of Chemistry, Cleveland State University, Ohio 44195, USA
    Biochemistry 44:13082-90. 2005
    ....
  13. ncbi Incorporation of factor Va into prothrombinase is required for coordinated cleavage of prothrombin by factor Xa
    Michael A Bukys
    Department of Chemistry, Cleveland State University, Cleveland, Ohio 44115, USA
    J Biol Chem 280:27393-401. 2005
    ..These data indicate that the interaction of factor Xa with the heavy chain of factor Va strongly influences the catalytic activity of the enzyme resulting in increased rates for both prothrombin-activating cleavages...
  14. ncbi Coagulation factor V: a plethora of anticoagulant molecules
    Michael Kalafatis
    Department of Chemistry, Cleveland State University, Cleveland, OH 44114, USA
    Curr Opin Hematol 12:141-8. 2005
    ..In this review, we summarize the current state of knowledge with respect to the interactions of the factor Va molecule with the various components of prothrombinase...