scholarly journals The contributions of Ca2+, phospholipids and tissue-factor apoprotein to the activation of human blood-coagulation factor X by activated factor VII

1990 ◽  
Vol 265 (2) ◽  
pp. 327-336 ◽  
Author(s):  
V J J Bom ◽  
R M Bertina
Keyword(s):  
Tissue Factor ◽  
Blood Coagulation ◽  
Reaction Rate ◽  
Factor Viia ◽  
Coagulation Factor ◽  
Fluid Phase ◽  
Fold Increase ◽  
Factor Vii ◽  
Factor X ◽  
Extrinsic Pathway

In the extrinsic pathway of blood coagulation, Factor X is activated by a complex of tissue factor, factor VII(a) and Ca2+ ions. Using purified human coagulation factors and a sensitive spectrophotometric assay for Factor Xa, we could demonstrate activation of Factor X by Factor VIIa in the absence of tissue-factor apoprotein, phospholipids and Ca2+. This finding allowed a kinetic analysis of the contribution of each of the cofactors. Ca2+ stimulated the reaction rate 10-fold at an optimum of 6 mM (Vmax. of 1.1 x 10(-3) min-1) mainly by decreasing the Km of Factor X (to 11.4 microM). In the presence of Ca2+, 25 microM-phospholipid caused a 150-fold decrease of the apparent Km and a 2-fold increase of the apparent Vmax. of the reaction; however, both kinetic parameters increased with increasing phospholipid concentration. Tissue-factor apoprotein contributed to the reaction rate mainly by an increase of the Vmax., in both the presence (40,500-fold) and absence (4900-fold) of phospholipid. The formation of a ternary complex of Factor VIIa with tissue-factor apoprotein and phospholipid was responsible for a 15 million-fold increase in the catalytic efficiency of Factor X activation. The presence of Ca2+ was absolutely required for the stimulatory effects of phospholipid and apoprotein. The data fit a general model in which the Ca2(+)-dependent conformation allows Factor VIIa to bind tissue-factor apoprotein and/or a negatively charged phospholipid surface resulting into a decreased intrinsic Km and an increased Vmax. for the activation of fluid-phase Factor X.

scholarly journals A candidate activation pathway for coagulation factor VII

2019 ◽  
Vol 476 (19) ◽  
pp. 2909-2926
Author(s):  
Tina M. Misenheimer ◽  
Kraig T. Kumfer ◽  
Barbara E. Bates ◽  
Emily R. Nettesheim ◽  
Bradford S. Schwartz
Keyword(s):  
Tissue Factor ◽  
Blood Coagulation ◽  
Factor Viia ◽  
Coagulation Factor ◽  
Factor Ix ◽  
Factor Vii ◽  
Factor X ◽  
Contact Activation ◽  
Coagulation Factor Vii ◽  
Factor Viiia

Abstract The mechanism of generation of factor VIIa, considered the initiating protease in the tissue factor-initiated extrinsic limb of blood coagulation, is obscure. Decreased levels of plasma VIIa in individuals with congenital factor IX deficiency suggest that generation of VIIa is dependent on an activation product of factor IX. Factor VIIa activates IX to IXa by a two-step removal of the activation peptide with cleavages occurring after R191 and R226. Factor IXaα, however, is IX cleaved only after R226, and not after R191. We tested the hypothesis that IXaα activates VII with mutant IX that could be cleaved only at R226 and thus generate only IXaα upon activation. Factor IXaα demonstrated 1.6% the coagulant activity of IXa in a contact activation-based assay of the intrinsic activation limb and was less efficient than IXa at activating factor X in the presence of factor VIIIa. However, IXaα and IXa had indistinguishable amidolytic activity, and, strikingly, both catalyzed the cleavage required to convert VII to VIIa with indistinguishable kinetic parameters that were augmented by phospholipids, but not by factor VIIIa or tissue factor. We propose that IXa and IXaα participate in a pathway of reciprocal activation of VII and IX that does not require a protein cofactor. Since both VIIa and activated IX are equally plausible as the initiating protease for the extrinsic limb of blood coagulation, it might be appropriate to illustrate this key step of hemostasis as currently being unknown.

Self-Damping Mechanism in Blood Coagulation

1974 ◽  
Vol 32 (01) ◽  
pp. 057-064 ◽  
Author(s):  
Y Nemerson ◽  
S.A Silverberg ◽  
J Jesty
Keyword(s):  
Tissue Factor ◽  
Blood Coagulation ◽  
Calcium Ions ◽  
Control Mechanism ◽  
Factor Vii ◽  
Damping Factor ◽  
Factor V ◽  
Factor X ◽  
Extrinsic Pathway ◽  
Two Systems

SummaryTwo reactions of the extrinsic pathway of coagulation, the activations of Factor X and prothrombin, have been studied in purified systems and shown to be self-damping. Factor X was activated by the tissue factor - Factor VII complex, and prothrombin by two systems: the coagulant protein of Taipan venom, and the physiological complex of activated Factor X, Factor V, lipid, and calcium ions. In each case the yield of enzyme, activated Factor X or thrombin, is a function of the concentration of activator. These and other observations are considered as a basis for a control mechanism in coagulation.

scholarly journals Studies of a mechanism inhibiting the initiation of the extrinsic pathway of coagulation

Blood ◽  
1987 ◽  
Vol 69 (2) ◽  
pp. 645-651 ◽  
Author(s):  
LV Rao ◽  
SI Rapaport
Keyword(s):  
Tissue Factor ◽  
Factor Viia ◽  
Factor Xa ◽  
Factor Ix ◽  
Factor Vii ◽  
Factor X ◽  
Extrinsic Pathway ◽  
Mechanism Of Inhibition ◽  
Peptide Release ◽  
Release Assay

Abstract We have extended earlier studies (Blood 66:204, 1985) of a mechanism of inhibition of factor VIIa/tissue factor activity that requires a plasma component (called herein extrinsic pathway inhibitor or EPI) and factor Xa. An activated peptide release assay using 3H-factor IX as a substrate was used to evaluate inhibition. Increasing the tissue factor concentration from 20% to 40% (vol/vol) overcame the inhibitory mechanism in normal plasma but not in factor VII-deficient plasma supplemented with a low concentration of factor VII. A second wave of factor IX activation obtained by a second addition of tissue factor to plasma with a normal factor VII concentration was almost abolished by supplementing the reaction mixture with additional EPI and factor X. Factor Xa's active site was necessary for factor Xa's contribution to inhibition, but preliminary incubation of factor Xa with EPI in the absence of factor VIIa/tissue factor complex or of factor VIIa/tissue factor complex in the absence of EPI did not replace the need for the simultaneous presence of factor Xa, factor VIIa/tissue factor, calcium, and EPI in an inhibitory reaction mixture. Inhibition of factor VIIa/tissue factor was reversible; both tissue factor and factor VIIa activity could be recovered from a dissociated, inhibited factor VIIa/tissue factor complex. EPI appeared to bind to a factor VIIa/tissue factor complex formed in the presence of factor Xa but not to a factor VIIa/tissue factor complex formed in the absence of factor Xa.

scholarly journals Studies of a mechanism inhibiting the initiation of the extrinsic pathway of coagulation

Blood ◽  
1987 ◽  
Vol 69 (2) ◽  
pp. 645-651 ◽  
Author(s):  
LV Rao ◽  
SI Rapaport
Keyword(s):  
Tissue Factor ◽  
Factor Viia ◽  
Factor Xa ◽  
Factor Ix ◽  
Factor Vii ◽  
Factor X ◽  
Extrinsic Pathway ◽  
Mechanism Of Inhibition ◽  
Peptide Release ◽  
Release Assay

We have extended earlier studies (Blood 66:204, 1985) of a mechanism of inhibition of factor VIIa/tissue factor activity that requires a plasma component (called herein extrinsic pathway inhibitor or EPI) and factor Xa. An activated peptide release assay using 3H-factor IX as a substrate was used to evaluate inhibition. Increasing the tissue factor concentration from 20% to 40% (vol/vol) overcame the inhibitory mechanism in normal plasma but not in factor VII-deficient plasma supplemented with a low concentration of factor VII. A second wave of factor IX activation obtained by a second addition of tissue factor to plasma with a normal factor VII concentration was almost abolished by supplementing the reaction mixture with additional EPI and factor X. Factor Xa's active site was necessary for factor Xa's contribution to inhibition, but preliminary incubation of factor Xa with EPI in the absence of factor VIIa/tissue factor complex or of factor VIIa/tissue factor complex in the absence of EPI did not replace the need for the simultaneous presence of factor Xa, factor VIIa/tissue factor, calcium, and EPI in an inhibitory reaction mixture. Inhibition of factor VIIa/tissue factor was reversible; both tissue factor and factor VIIa activity could be recovered from a dissociated, inhibited factor VIIa/tissue factor complex. EPI appeared to bind to a factor VIIa/tissue factor complex formed in the presence of factor Xa but not to a factor VIIa/tissue factor complex formed in the absence of factor Xa.

Purification and Properties of an Abnormal Blood Coagulation Factor IX (Factor IXBm)/Kinetics of Its Inhibition of Factor X Activation by Factor VII and Bovine Tissue Factor

1981 ◽  
Vol 45 (01) ◽  
pp. 055-059 ◽  
Author(s):  
B Østerud ◽  
C K Kasper ◽  
K K Lavine ◽  
C Prodanos ◽  
S I Rapaport
Keyword(s):  
Tissue Factor ◽  
Blood Coagulation ◽  
Limited Proteolysis ◽  
Coagulation Factor ◽  
Factor Ix ◽  
Factor Vii ◽  
Factor X ◽  
Factor Xi ◽  
Coagulation Factor Ix ◽  
Bovine Tissue

SummaryAn abnormal blood coagulation factor IX has been isolated from the blood of a hemophilia B patient with a variant of the disease (hemophilia Bm) characterized by a normal concentration of factor IX antigen, negligible factor IX coagulant activity, and a prolonged prothrombin time with bovine tissue factor. The isolated protein (factor IXBm) had the same apparent molecular weight as normal factor IX (55,000) and the same mobility on two dimensional immunoelectrophoresis as normal factor IX. Factor IXBm underwent limited proteolysis induced by activated factor XI, in the presence of Ca2+ ions, or induced by the reaction product of tissue factor, factor VII and Ca2+ ions. A timecourse study showed that activated factor XI cleaved factor IXBm and factor IX at similar rates. However, in contrast to normal factor IX, the limited protelysis of factor IXBm did not generate procoagulant activity.In kinetic experiments purified factor IXBm behaved like a competitive inhibitor (Ki of 0.017 μM) of the activation of factor X by bovine tissue factor and factor VII. Normal factor IX was also found to inhibit the reaction but required a four-fold higher concentration to achieve the same inhibitory effects as factor IXBm.

scholarly journals Role of Zymogen and Activated Factor X as Scaffolds for the Inhibition of the Blood Coagulation Factor VIIa-Tissue Factor Complex by Recombinant Nematode Anticoagulant Protein c2

2001 ◽  
Vol 276 (13) ◽  
pp. 10063-10071 ◽  
Author(s):  
Peter W. Bergum ◽  
Amy Cruikshank ◽  
Steven L. Maki ◽  
Curtis R. Kelly ◽  
Wolfram Ruf ◽  
...  
Keyword(s):  
Tissue Factor ◽  
Blood Coagulation ◽  
Factor Viia ◽  
Coagulation Factor ◽  
Factor X

scholarly journals Kinetics of Factor X Activation Catalyzed by Factor VII.

1977 ◽  
Author(s):  
Yale Nemerson
Keyword(s):  
Tissue Factor ◽  
Factor Viia ◽  
Normal Population ◽  
Fold Increase ◽  
Factor Vii ◽  
Factor X ◽  
Single Polypeptide Chain ◽  
Upward Deflection ◽  
Single Polypeptide ◽  
Factor X Activation

Factor X activation is the point at which the intrinsic and extrinsic pathways of coagulation converge. Accordingly, events which control the rate of activation of Factor X might be central to the overall control of coagulation. We have therefore attempted to extract the kinetic parameters which govern this reaction. We have also devised and utilized a new assay for this reaction based on the release of radiolabeled activation peptide from Factor X. Utilizing Factor VIIa (2-chain VII) in the presence of saturating amounts of tissue factor, the KM was found to be 0.34 μ M and the Kcat 32 sec-1. The value for the KM is about twice the concentration of Tactor X in plasma. This means that rather small variations in Factor X concentration, such as those encountered in the normal population, would have significant effects on the rate at which Factor X is activated. We have also found that the effect of tissue factor on this reaction is mainly on the catalytic activity of Factor VII although in the absence of tissue factor there is also a 10-fold increase in the KM. Factor VII is predominantly present in plasma as a single polypeptide chain. Utilizing similar techniques we have also examined the reaction catalyzed by this form of Factor VII. All progress curves showed a prolonged lag followed by a sharp upward deflection. The mechanism underlying this phenomenon is now being investigated and will be discussed.

Blocking the Initiation of Coagulation by RNA Aptamers to Factor VIIa

2000 ◽  
Vol 84 (11) ◽  
pp. 841-848 ◽  
Author(s):  
H. Lyerly ◽  
Jeffrey Lawson ◽  
Christopher Rusconi ◽  
Alice Yeh ◽  
Bruce Sullenger
Keyword(s):  
Human Plasma ◽  
Tissue Factor ◽  
In Vitro Selection ◽  
Factor Viia ◽  
Coagulation Factor ◽  
Factor Vii ◽  
Dependent Manner ◽  
Factor X ◽  
Coagulation Factor Vii

SummaryThe tissue factor/factor VIIa complex is thought to be the primary initiator of most physiologic blood coagulation events. Because of its proximal role in this process, we sought to generate new inhibitors of tissue factor/factor VIIa activity by targeting factor VIIa. We employed a combinatorial RNA library and in vitro selection methods to isolate a high affinity, nuclease-resistant RNA ligand that binds specifically to coagulation factor VII/VIIa. This RNA inhibits the tissue factordependent activation of factor X by factor VIIa. Kinetic analyses of the mechanism of action of this RNA suggest that it antagonizes factor VIIa activity by preventing formation of a functional factor VII/tissue factor complex. Furthermore, this RNA significantly prolongs the prothrombin time of human plasma in a dose dependent manner, and has an in vitro half-life of ∼15 h in human plasma. Thus, this RNA ligand represents a novel class of anticoagulant agents directed against factor VIIa.

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