PDF herunterladen
Thromb Haemost 1985; 53(03): 396-400
DOI: 10.1055/s-0038-1661322
Original Article
Schattauer GmbH Stuttgart

Assembly of the Intrinsic Factor X Activating Complex – Interactions between Factor IXa, Factor VIIIa and Phospholipid

Gerbrand van Dieijen
The Department of Biochemistry, University of Limburg, Maastricht, The Netherlands
,
Jan L M L van Rijn
The Department of Biochemistry, University of Limburg, Maastricht, The Netherlands
,
José W P Govers-Riemslag
The Department of Biochemistry, University of Limburg, Maastricht, The Netherlands
,
H Coenraad Hemker
The Department of Biochemistry, University of Limburg, Maastricht, The Netherlands
,
Jan Rosing
The Department of Biochemistry, University of Limburg, Maastricht, The Netherlands
› Institutsangaben
Weitere Informationen

Publikationsverlauf

Received 05. November 1984

Accepted 22. März 1985

Publikationsdatum:
18. Juli 2018 (online)

Auch verfügbar auf
  als PDF herunterladen  Lizenzen und Reprints

Summary

The activation of blood coagulation factor X by factor IXa is strongly stimulated by the non-enzymatic cofactors phospholipid, Ca2+ and activated factor VIII. In this paper we present a method by which we were able to determine binding affinities of factor IXa for phospholipids (either in the absence or presence of factor VIIIa) from kinetic measurements of factor X activation. It is shown that rates of factor X activation in the presence of phospholipids can be saturated with an excess factor VIIIa at limiting amounts of factor IXa and vice versa. Our data indicate that the enzymatic unit in the intrinsic factor X activator is a 1:1 stoichiometrical complex of factor IXa and factor VIIIa bound to phospholipid. Titrations with factor IXa at fixed concentrations of phospholipid and factor X show that the apparent dissociation constant of factor IXa for phospholipid is lowered from 10-6 M to 10-8 M by the presence of factor VIIIa. We conclude, that in analogy with the role of factor Va in prothrombin activation, phospholipid-bound factor VIIIa functions as a high-affinity binding site (»receptor«) for factor IXa in the intrinsic factor X activating complex. Therefore, factor VIIIa increases the observed Vmax of factor X activation by 1) enhancing the kcat of the reaction and 2) increasing the amount of phospholipid-bound factor IXa that participates in factor X activation.

Keywords

Intrinsic factor X activation - Factor VIII - Kinetics and mechanism - Binding parameters
 

  • References

  • 1 Jackson CM, Nemerson Y. Blood coagulation Ann Rev Biochem. 1980; 49: 765-811
    PubMedSuche in Google Scholar
  • 2 Hoyer LW. The factor VIII complex: structure and function. Blood 1981; 58: 1-13
    PubMedSuche in Google Scholar
  • 3 Hemker HC, Kahn MJ P. Reaction sequence of blood coagulation. Nature 1967; 215: 1201-1202
    CrossrefPubMedSuche in Google Scholar
  • 4 Østerud B, Rapaport SI. Synthesis of intrinsic factor X activator. Inhibition of the function of formed activator by antibodies to factor VIII and to factor IX Biochemistry 1970; 9: 1854-1861
    CrossrefPubMedSuche in Google Scholar
  • 5 Hultin MB, Nemerson Y. Activation of factor X by factors IXa and VIII; a specific assay for factor IXa in the presence of thrombin activated factor VIII. Blood 1978; 52: 928-940
    PubMedSuche in Google Scholar
  • 6 van Dieijen G, Tans G, Rosing J, Hemker HC. The role of phospholipid and factor VIIIa in the activation of bovine factor X. J Biol Chem 1981; 256: 3433-3442
    Suche in Google Scholar
  • 7 Griffith MJ, Reisner HM, Lundblad RL, Roberts HR. Measurement of human factor IXa activity in an isolated factor X activation system. Thromb Res 1982; 27: 289-301
    CrossrefPubMedSuche in Google Scholar
  • 8 Nelsestuen GL, Kiesel W, DiScipio RG. Interaction of vitamin K dependent proteins with membranes. Biochemistry 1978; 17: 2134-2138
    CrossrefPubMedSuche in Google Scholar
  • 9 van Dieijen G, Tans G, van Rijn JL M L, Zwaal RF A, Rosing J. Simple and rapid method to determine the binding of blood clotting factor X to phospholipid vesicles. Biochemistry 1981; 20: 7096-7101
    CrossrefPubMedSuche in Google Scholar
  • 10 Rosing J, Tans G, Govers-Riemslag JW P, Zwaal RF A, Hemker HC. The role of phospholipids and factor Va in the prothrombinase complex. J Biol Chem 1980; 255: 274-283
    PubMedSuche in Google Scholar
  • 11 Nesheim ME, Taswell JB, Mann KG. The contribution of bovine factor V and factor Va to the activity of prothrombinase. J Biol Chem 1979; 254: 10952-10962
    PubMedSuche in Google Scholar
  • 12 Lindhout T, Govers-Riemslag JW P, van de Waart P, Hemker HC, Rosing J. Factor Va-factor Xa interaction. Effects of phospholipid vesicles of varying composition Biochemistry 1982; 21: 5494-5502
    CrossrefPubMedSuche in Google Scholar
  • 13 Comfurius P, Zwaal RF A. The enzymatic synthesis of phosphatidylserine and purification by CM-cellulose column chromatography. Biochim Biophys Acta 1977; 488: 36-42
    CrossrefPubMedSuche in Google Scholar
  • 14 Vehar GA, Davie EW. Preparation and properties of bovine factor VIII. Biochemistry 1980; 19: 401-410
    CrossrefPubMedSuche in Google Scholar
  • 15 Tans G, Janssen-Claessen T, van Dieijen G, Hemker HC, Rosing J. Activation of factor IX by factor XIa . A spectrophotometric assay for factor IX in human plasma Thromb Haemostas 1982; 48: 127-132
    PubMedSuche in Google Scholar
  • 16 DiScipio RG, Hermodson MA, Yates SG, Davie EW. A comparison of human prothrombin, factor IX, factor X, and Protein S. Biochemistry 1977; 16: 698-706
    CrossrefPubMedSuche in Google Scholar