Polyphosphate binds with high affinity to exosite II of thrombin
N J Mutch1, T Myles, L L K Leung
1Department of Biochemistry, College of Medicine, University of Illinois at Urbana-Champaign, Urbana, IL 61801, USA.
Journal of Thrombosis and Haemostasis : JTH
|December 17, 2009
Summary
Polyphosphate, released by platelets, binds to thrombin's exosite II. This interaction, with a low dissociation constant, may be significant in blood coagulation.
Area of Science:
- Biochemistry
- Hematology
- Molecular Biology
Background:
- Polyphosphate is secreted from platelet dense granules.
- Polyphosphate accelerates factor V activation by thrombin.
Purpose of the Study:
- To examine the interaction of polyphosphate with thrombin.
Main Methods:
- Gel mobility assays to assess polyphosphate-thrombin interaction.
- Mutagenesis of thrombin exosites.
- Surface plasmon resonance (SPR) for binding kinetics.
- Competition assays with glycosaminoglycans.
Main Results:
- Thrombin binds polyphosphate, primarily through exosite II.
- SPR revealed a tight interaction (K(d) ≈ 5 nM).
- Polyphosphate binding partially overlaps with, but is distinct from, the heparin-binding site.
Conclusions:
- Polyphosphate interacts with thrombin via exosite II.
- This interaction is likely physiologically relevant due to achievable polyphosphate concentrations in vivo.
- Polyphosphate does not interfere with heparin's anticoagulant function via antithrombin.
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