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Thromb Haemost 2010; 103(02): 291-298
DOI: 10.1160/TH09-07-0420
Blood Coagulation, Fibrinolysis and Cellular Haemostasis
Schattauer GmbH

Proteolytic resistance conferred to fibrinogen by von Willebrand factor

Anna Tanka-Salamon
1   Department of Medical Biochemistry, Semmelweis University, Budapest, Hungary
,
Krasimir Kolev
1   Department of Medical Biochemistry, Semmelweis University, Budapest, Hungary
,
Raymund Machovich
1   Department of Medical Biochemistry, Semmelweis University, Budapest, Hungary
,
Erzsebet Komorowicz
1   Department of Medical Biochemistry, Semmelweis University, Budapest, Hungary
› Author Affiliations Financial support: This work was supported by the Hungarian Scientific Research Fund [OTKA-F42475], [OTKA 75430], [OTKA K60123], Medical Scientific Council [ETT 005/2009] and the Wellcome Trust [083174/B/07/Z].
Further Information

Publication History

Received: 01 July 2009

Accepted after major revision: 05 November 2009

Publication Date:
22 November 2017 (online)

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Summary

The formation of platelet-rich thrombi under high shear rates requires both fibrinogen and von Willebrand factor (VWF) as molecular adhesives between platelets. We attempted to describe the role of VWF as a potential substrate and modulator of the fibrinolytic system using binding assays, as well as kinetic measurements on the cleavage of fibrin(ogen) and a synthetic plasmin substrate (Spectrozyme-PL). The similar dissociation constants for the binding of plasminogen, plasmin, and active site-blocked plasmin onto immobilised VWF suggest that the primary binding site in plasmin(ogen) is not the active site. The progressive loss of clottability and generation of degradation products during fibrinogen digestion with plasmin were delayed in the presence of VWF at physiological concentrations, while VWF cleavage was not detectable. Determination of kinetic parameters for fibrinogen degradation by plasmin, miniplasmin and microplasmin showed that VWF did not modify the Km, whereas kcat values decreased with increasing VWF concentrations following the kinetic model of non-competitive inhibition. Inhibitory constants calculated for VWF were in the range of its physiological plasma concentration (5.4 μg/ml, 5.7 μg/ml and 10.0 μg/ ml for plasmin, miniplasmin and microplasmin, respectively) and their values suggested a modulating role of the kringle 5 domain in the interaction between VWF and (mini)plasmin. VWF had no effect on the amidolytic activity of plasmin on Spectrozyme-PL, or on fibrin dissolution by (mini)plasmin. Our data suggest that VWF, while a poor plasmin substrate relative to fibrinogen, protects fibrinogen against degradation by plasmin preserving its clottability in plasma and its adhesive role in platelet-rich thrombi.

Keywords

von Willebrand factor - fibrinogen - plasmin - fibrinolysis - thrombolysis
 

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