The mechanism of VWF-mediated platelet GPIbalpha binding.
The binding of Von Willebrand Factor to platelets is dependent on the conformation of the A1 domain which binds to platelet GPIbalpha. This interaction initiates the adherence of platelets to the subendothelial vasculature under the high shear that occurs in pathological thrombosis. We have developed a thermodynamic strategy that defines the A1:GPIbalpha interaction in terms of the free energies (DeltaG values) of A1 unfolding from the native to intermediate state and the binding of these conformational states to GPIbalpha. We have isolated the intermediate conformation of A1 under nondenaturing conditions by reduction and carboxyamidation of the disulfide bond. The circular dichroism spectrum of reduction and carboxyamidation A1 indicates that the intermediate has approximately 10% less alpha-helical structure that the native conformation. The loss of alpha-helical secondary structure increases the GPIbalpha binding affinity of the A1 domain approximately 20-fold relative to the native conformation. Knowledge of these DeltaG values illustrates that the A1:GPIbalpha complex exists in equilibrium between these two thermodynamically distinct conformations. Using this thermodynamic foundation, we have developed a quantitative allosteric model of the force-dependent catch-to-slip bonding that occurs between Von Willebrand Factor and platelets under elevated shear stress. Forced dissociation of GPIbalpha from A1 shifts the equilibrium from the low affinity native conformation to the high affinity intermediate conformation. Our results demonstrate that A1 binding to GPIbalpha is thermodynamically coupled to A1 unfolding and catch-to-slip bonding is a manifestation of this coupling. Our analysis unites thermodynamics of protein unfolding and conformation-specific binding with the force dependence of biological catch bonds and it encompasses the effects of two subtypes of mutations that cause Von Willebrand Disease.
Auton
Matthew
M
Cardiovascular Research, Baylor College of Medicine, Houston, Texas, USA.
Zhu
Cheng
C
Cruz
Miguel A
MA
D000818:Animals; D001696:Biomechanical Phenomena; D001792:Blood Platelets / Q000378:metabolism*; D002417:Cattle; D002850:Chromatography, Gel; D007700:Kinetics; D019038:Platelet Glycoprotein GPIb-IX Complex / Q000378:metabolism*; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D012710:Serum Albumin, Bovine / Q000378:metabolism; D013816:Thermodynamics; D014841:von Willebrand Factor / Q000378:metabolism*
D019038:Platelet Glycoprotein GPIb-IX Complex; D014841:von Willebrand Factor; D012710:Serum Albumin, Bovine