The activation of factor V by factor Xa or α-chymotrypsin and comparison with thrombin and RVV-V action. An improved factor V isolation procedure

Smith, Catherine M.; Hanahan, Donald J.

doi:10.1021/bi00654a007

Cited by 44 publications

(20 citation statements)

References 28 publications

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“…We observed that for the expression of amidolytic activity, RVV‐V and LVV‐V do not require a functional calcium binding site, since both FV activators retained amidase activity after treatment with the metal chelator EDTA (Table I). Furthermore, it has been demonstrated that both snake venom proteases activate FV in a calcium‐independent manner 17, 67…”

Section: Resultsmentioning

confidence: 99%

Structural models of the snake venom factor V activators from Daboia russelli and Daboia lebetina

Segers¹,

Rosing²,

Nicolaes³

2006

Proteins

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Blood coagulation factor V (FV) is a multifunctional protein that circulates in human plasma as a precursor molecule which can be activated by thrombin or activated factor X (FXa) in order to express its cofactor activity in prothrombin activation. FV activation is achieved by limited proteolysis after Arg709, Arg1018, and Arg1545 in the FV molecule. The venoms of Daboia russelli and Daboia lebetina contain a serine protease that specifically activates FV by a single cleavage at Arg1545. We have predicted the three-dimensional structure of these enzymes using comparative protein modeling techniques. The plasminogen activator from Agkistrodon acutus, which shows a high degree of homology with the venom FV activators and for which a high-quality crystallographic structure is available, was used as the molecular template. The RVV-V and LVV-V models provide for the first time a detailed and accurate structure of a snake venom FV activator and explain the observed sensitivity or resistance toward a number of serine protease inhibitors. Finally, electrostatic potential calculations show that two positively charged surface patches are present on opposite sides of the active site. We propose that both FV activators achieve their exquisite substrate specificity for the Arg1545 site via interactions between these exosites and FV.

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Section: Resultsmentioning

confidence: 99%

Structural models of the snake venom factor V activators from Daboia russelli and Daboia lebetina

Segers¹,

Rosing²,

Nicolaes³

2006

Proteins

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“…Bovine Factor V is more stable than its human counterpart, for which reason most of our knowledge derives from studies using bovine material (9)(10)(11)(12)(13)(14). However, only recently was bovine Factor V purified to Received for publication 26 February 1980 and in revised form 2 May 1980. homogeneity, by Nesheim et al (12) and by Esmon (13).…”

Section: Introductionmentioning

confidence: 99%

Human coagluation factor V purification and thrombin-catalyzed activation.

Dahlbäck¹

1980

J. Clin. Invest.

150

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A B S T R A C T Factor V was isolated from human plasma by barium citrate adsorption, polyethylene glycol fractionation, DEAE-Sepharose CL-6B chromatography, ammonium sulfate fractionation, and gel chromatography on Ultrogel 22. Degradation of Factor V during purification was largely prevented by ample use of inhibitors of proteolytic enzyme. The purified Factor V was a stable, single-chain molecule with an apparent molecular weight of 330,000. Activation of human Factor V by thrombin resulted in a 10-to 15-fold increase in activity. The activation pattem as monitored by sodium dodecyl sulfate polyacrylamide gel electrophoresis was compared with that of bovine Factor V. Differences in the patterns of thrombin activation were noticed between the two species, whereas the final products were similar. The products ofhuman Factor V activation are two closely spaced doublets, one with an apparent molecular weight of -110,000, and the other, -72,000. An antibody was raised against the purified protein. Crossed immunoelectrophoresis showed that the antibody recognized Factor V both before and after activation with thrombin.

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“…Tissue factor pathway inhibitor (TFPI) binds to TF-FVIIa-FXa to limit the production of FXa and FIXa by TF-FVIIa (3,4). Nevertheless, once produced, thrombin and the initially formed FXa activate small quantities of factor V (FV) to FVa and factor VIII (FVIII) to FVIIIa (5)(6)(7)(8). The activation of these two cofactors leads to the formation of two other essential procoagulant complexes, both involving FX, at the surface of procoagulant phospholipids in the presence of calcium ions (9), FIXa-FVIIIa and FXa-FVa complexes, which convert FX to FXa and prothrombin to thrombin, respectively.…”

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confidence: 99%

Role of the Gla and First Epidermal Growth Factor-like Domains of Factor X in the Prothrombinase and Tissue Factor-Factor VIIa Complexes

Thiec

Cherel

Olivier

2003

Journal of Biological Chemistry

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Factor X (FX) has high structure homology with other proteins of blood coagulation such as factor IX (FIX) and factor VII (FVII). These proteins present at their aminoterminal extremity a ␥-carboxyglutamic acid containing domain (Gla domain), followed by two epidermal growth factor-like (EGF1 and EGF2) domains, an activation peptide, and a serine protease domain. After vascular damage, the tissue factor-FVIIa (TF-FVIIa) complex activates both FX and FIX. FXa interacts stoichiometrically with tissue pathway inhibitor (TFPI), regulating TF-FVIIa activity by forming the TF-FVIIa-TFPI-FXa quaternary complex. Conversely, FXa boosts coagulation by its association with its cofactor, factor Va (FVa). To investigate the contribution of the Gla and EGF1 domains of FX in these complexes, FX chimeras were produced in which FIX Gla and EGF1 domains substituted the corresponding domains of FX. The affinity of the two chimeras, FX/FIX(Gla) and FX/FIX(EGF1), for the TF-FVIIa complex was markedly reduced compared with that of wild-type-FX (wt-FX) independently of the presence of phospholipids. Furthermore, the association rate constants of preformed FX/FIX(Gla)-TFPI and FX/FIX(EGF1)-TFPI complexes with TF-FVIIa were, respectively, 10-and 5-fold slower than that of wt-FXa-TFPI complex. Finally, the apparent affinity of FVa was 2-fold higher for the chimeras than for wt-FX in the presence of phospholipids and equal in their absence. These data demonstrate that FX Gla and EGF1 domains contain residues, which interact with TF-FVIIa exosites contributing to the formation of the TF-FVIIa-FX and TF-FVIIa-TFPI-FXa complexes. On the opposite, FXa Gla and EGF1 domains are not directly involved in FVa binding.The blood coagulation cascade consists of a series of enzymatic conversions driven by the formation of complexes between serine proteases and cell membrane-bound cofactors. Human factor X (FX) 1 is one of the serine protease zymogens playing a central role in coagulation processes leading to the formation of a fibrin clot. This is illustrated by the behavior of FX as a substrate or as an enzyme in three essential blood coagulation complexes. First, FX is a natural substrate, as well as factor IX (FIX), of the tissue factor-factor VIIa (TF-FVIIa) complex (1) considered as the initial enzyme complex in the cascade following vascular damage. FX activation by TF-FVIIa results from specific cleavage and release of a 52-residue activation peptide. Activated FX (FXa) can generate a tiny amount of thrombin from prothrombin in an extremely inefficient reaction (2). Tissue factor pathway inhibitor (TFPI) binds to TF-FVIIa-FXa to limit the production of FXa and FIXa by TF-FVIIa (3, 4). Nevertheless, once produced, thrombin and the initially formed FXa activate small quantities of factor V (FV) to FVa and factor VIII (FVIII) to FVIIIa (5-8). The activation of these two cofactors leads to the formation of two other essential procoagulant complexes, both involving FX, at the surface of procoagulant phospholipids in the presence of calcium ions (9),...

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The activation of factor V by factor Xa or α-chymotrypsin and comparison with thrombin and RVV-V action. An improved factor V isolation procedure

Cited by 44 publications

References 28 publications

Structural models of the snake venom factor V activators from Daboia russelli and Daboia lebetina

Structural models of the snake venom factor V activators from Daboia russelli and Daboia lebetina

Human coagluation factor V purification and thrombin-catalyzed activation.

Role of the Gla and First Epidermal Growth Factor-like Domains of Factor X in the Prothrombinase and Tissue Factor-Factor VIIa Complexes

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