Purified A2 domain of von Willebrand factor binds to the active conformation of von Willebrand factor and blocks the interaction with platelet glycoprotein Ibalpha.

Abstract

BACKGROUND:

von Willebrand factor (VWF) does not interact with circulating platelets unless it is induced to expose the binding site for platelet glycoprotein (GP)Ibalpha in the A1 domain by high shear stress, immobilization, and/or a modulator. Previous studies have implied indirectly that the A2 domain may be involved in regulating A1-GPIbalpha binding.

OBJECTIVE AND METHODS:

Because the relationship between the A1 and A2 domains has not been defined, we have investigated the effect of the A2 domain on the binding activity of the A1 domain using recombinant A domain polypeptides, multimeric VWF, and monoclonal antibodies (mAb).

RESULTS:

The A2 domain polypeptide bound specifically to the immobilized A1 domain polypeptide or full-length VWF, with half-maximal binding being obtained at 60 or 168 nm, respectively. This A1-A2 interaction was inhibited by mAb against the A2 or A1 domain and by the A1 domain polypeptide. The A2 domain polypeptide effectively blocked GPIbalpha-mediated platelet adhesion under high flow conditions. The A2 domain polypeptide specifically recognizes the GPIbalpha-binding conformation in the A1 domain, as it only interacted with VWF activated by the modulator ristocetin or immobilized VWF. Furthermore, in contrast to plasma VWF, the ultra-large (UL)VWF multimers or a recombinant VWF-A1A2A3 polypeptide containing a gain-of-function mutation (R1308 L) of type 2B von Willebrand disease bound to the A2 domain polypeptide without the need for ristocetin.

CONCLUSIONS:

The recombinant A2 domain polypeptide specifically binds to the active conformation of the A1 domain in VWF and effectively blocks the interaction with platelet GPIbalpha under high-flow conditions.

• Authors: C Martin, L D Morales, M A Cruz
• Journal: Journal of thrombosis and haemostasis : JTH (J Thromb Haemost) Vol. 5 Issue 7 Pg. 1363-70 (Jul 2007) ISSN: 1538-7933 [Print] England
• PMID: 17389010 (Publication Type: Journal Article, Research Support, N.I.H., Extramural)

Chemical References

• DNA Primers
• Platelet Glycoprotein GPIb-IX Complex
• Recombinant Proteins
• von Willebrand Factor

Topics

• Base Sequence
• Blood Platelets (metabolism)
• DNA Primers (genetics)
• Hemorheology
• Humans
• In Vitro Techniques
• Mutagenesis, Site-Directed
• Platelet Adhesiveness (physiology)
• Platelet Glycoprotein GPIb-IX Complex (metabolism)
• Protein Binding
• Protein Conformation
• Protein Structure, Quaternary
• Protein Structure, Tertiary
• Recombinant Proteins (chemistry, genetics, metabolism)
• von Willebrand Factor (chemistry, genetics, metabolism)