Interaction of thrombin and antithrombin. Reaction observed by intrinsic fluorescence measurements

Wong, Raymond F.; Windwer, Steven R.; Feinman, Richard D.

doi:10.1021/bi00286a001

Cited by 20 publications

(6 citation statements)

References 31 publications

(35 reference statements)

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“…RCL is inserted as strand 4 in 6-stranded bsheet A. much slower. Typical uncatalyzed in vitro thrombin inhibition rates at pH 7.4 and 25 C lie in the range of 7-11 Â 10 3 M À1 sec À1 , [31][32][33] whereas those for factor Xa are 2-3 Â 10 3 M À1 sec À1 . 34 Thus, the uncatalyzed reaction of antithrombin does not contribute much under physiological conditions.…”

Section: R a T E S O F A N T I T H R O M B I N I N H I B I T I O N mentioning

confidence: 99%

New antithrombin‐based anticoagulants

Desai

2003

Medicinal Research Reviews

131

115

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Clinically used anticoagulants are inhibitors of enzymes involved in the coagulation pathway, primarily thrombin and factor Xa. These agents can be either direct or indirect inhibitors of clotting enzymes. Heparin-based anticoagulants are indirect inhibitors that enhance the proteinase inhibitory activity of a natural anticoagulant, antithrombin. Despite its phenomenal success, current anticoagulation therapy suffers from the risk of serious bleeding. The need for safer and more effective antithrombotic agents clearly exists. The past decade has seen enormous effort directed toward discovering and/or designing new molecules with anticoagulant activity. These new molecules can be classified into (a). antithrombin and its mutants, (b). natural polysaccharides, (c). synthetic modified heparins and heparin-mimics, (d). synthetic oligosaccharides, and (e). synthetic non-sugar antithrombin activators. This review focuses on these efforts in designing or discovering new molecules that act through the antithrombin pathway of anticoagulation.

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Section: R a T E S O F A N T I T H R O M B I N I N H I B I T I O N mentioning

confidence: 99%

New antithrombin‐based anticoagulants

Desai

2003

Medicinal Research Reviews

131

115

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“…The inactivation of proteinases by antithrombin in the absence of heparin occurs rather slowly. The bimolecular rate constant for the reaction with thrombin around neutral pH is 7 × 10 3 to 1.1 × 10 4 M -1 s -1 at 25ºC [67][68][69][70] and approximateiy 1.4 × 104 M -1 s -1 at 37°C. 71 The latter value would correspond to a half-life of thrombin of ~20 s at 37°C under pseudo-first order conditions at plasma concentrations of antithrombin.…”

Section: Reaction With Thrombin and Other Clotting Proteinasesmentioning

confidence: 99%

“…This con clusion is indicated by the demonstration that the antithrombin-thrombin complex is not thermodynamically, but only kinetically, stable and slowly, with a half-life of about 3 days, dissociates to intact enzyme and reactive bond-cleaved inhibitor. 110,111 The changes in antigenic, spectroscopic, and heparin-binding properties of an tithrombin that accompany complex formation 69,[112][113][114][115][116] suggest that the trapping involves a conformational change of the inhibitor. Such a change is also consistent with the two-step mechanism, involving a stabilization of an initial weak antithrombin-thrombin complex, that has been demonstrated by rapid-kinetics studies.…”

Section: Reaction With Thrombin and Other Clotting Proteinasesmentioning

confidence: 99%

Regulation of Thrombin Activity by Antithrombin and Heparin

Olson¹,

Björk²

1994

Semin Thromb Hemost

136

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The central role of thrombin in blood clotting is well established. The classic function of thrombin in this pro cess is to cleave fibrinogen, a reaction that leads to for mation of the fibrin gel, the essential constituent of the blood clot. However, thrombin also has important regu latory functions in blood clotting. It thus activates the cofactors, Factor V and Factor VIII, of the clotting sys tem in a positive feedback loop that greatly accelerates the generation of the enzyme and thereby increases the rate of fibrin formation. In addition, thrombin can also retard blood clotting in a negative feedback loop. When bound to a membrane protein, thrombomodulin, on the endothelial cell surface, thrombin thus activates protein C, which in turn inactivates Factors V and VIII, thereby decreasing the rate of generation of thrombin and conse quently also of fibrin. A further function of thrombin is to activate Factor XIII, which then stabilizes the clot by covalent crosslinks. These divergent roles of thrombin in the clotting system suggest that precise control of the activity of the enzyme is of paramount importance for normal hemostasis.The activity of thrombin generated in the clotting process has been shown to be predominantly regulated by inactivation of the enzyme by four different plasma pro teinase inhibitors. Two of these, α 1 -proteinase inhibitor and α 2 -macroglobulin, are general proteinase inhibitors that can inactivate a number of enzymes, whereas the other two, antithrombin and heparin cofactor II, are more specific for thrombin and, in the case of antithrombin, also other coagulation proteinases. The latter two inhibi tors also have in common that the rate of their inactiva tion of thrombin is greatly accelerated by the binding of heparin and certain similar glycosaminoglycans.In this work we will review the structure and func tion of antithrombin, the physiologically most important of the two specific thrombin inhibitors. We will discuss the properties of antithrombin and how it inactivates thrombin and other clotting proteinases. We will also present current knowledge regarding the binding of hep arin to antithrombin and thrombin or other proteinases and the role of these interactions in the mechanism by which heparin accelerates the inhibition of the protein ases by the inhibitor. Finally, we will discuss the physio logical role of antithrombin and heparin-like polysaccha rides and how other proteins in plasma, on the vessel wall, or released by platelets and neutrophils may modu late the effect of heparin in accelerating proteinase inacti vation by antithrombin.

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“…Second-order rate constants were determined on an Apple 11+ microcomputer by using a non-linearregression analyis as described by Wong et al (1983). For thiol-group release, the rate constant for approach to the final slope is calculated.…”

Section: Kinetic Analysismentioning

confidence: 99%

Kinetics of the reaction of thrombin and α2-macroglobulin

Feinman¹,

Yuan²,

Windwer³

et al. 1985

Biochemical Journal

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The kinetics of the reaction of alpha 2-macroglobulin (alpha 2M) with human thrombin were studied by recording the appearance of thiol groups spectrophotometrically and by measuring the distribution of protein species by denaturing non-reducing gel electrophoresis. The goals were to study the relation between the formation of various covalent enzyme-inhibitor complex species and the appearance of free thiol, and from the kinetic analysis, to try to characterize the chemical nature of the protein complexes. The kinetics of thiol-group release were observed to be biphasic, the early phase showing second-order behaviour, results consistent with previous reports in the literature. The observed second-order rate constant for thiol-group release was found to be faster than the second-order rate constant for the disappearance of the band corresponding to native alpha 2M on gel electrophoresis. This may be a reflection of the multiple products formed from the thioester. Alternatively, it is possible that covalent-bond formation is slower than some enzyme-induced change in the thioester centre, and this may be suggestive evidence for a reactive alpha 2M centre that does not contain an intact thioester. The kinetics of covalent-bond formation were found to be consistent with the internal cross-link of several alpha 2M chains by the bound proteinase, providing further evidence that the very-high-Mr species seen on gels may arise from dimers of the alpha 2M molecule held together by covalent bonds to the enzyme.

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Interaction of thrombin and antithrombin. Reaction observed by intrinsic fluorescence measurements

Cited by 20 publications

References 31 publications

New antithrombin‐based anticoagulants

New antithrombin‐based anticoagulants

Regulation of Thrombin Activity by Antithrombin and Heparin

Kinetics of the reaction of thrombin and α2-macroglobulin

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