title: N-terminal flanking region of A1 domain in von Willebrand factor stabilizes structure of A1A2A3 complex and modulates platelet activation under shear stress.
abstract: von Willebrand factor (vWF) mediates platelet adhesion and thrombus formation via its interaction with the platelet receptor glycoprotein (GP)Ibα. We have analyzed two A1A2A3 tri-domain proteins to demonstrate that the amino acid sequence, Gln(1238)-Glu(1260), in the N-terminal flanking region of the A1 domain, together with the association between the A domains, modulates vWF-GPIbα binding and platelet activation under shear stress. Using circular dichroism spectroscopy and differential scanning calorimetry, we have described that sequence Gln(1238)-Glu(1260) stabilizes the structural conformation of the A1A2A3 tri-domain complex. The structural stabilization imparted by this particular region inhibits the binding capacity of the tri-domain protein for GPIbα. Deletion of this region causes a conformational change in the A1 domain that increases binding to GPIbα. Only the truncated protein was capable of effectively blocking ristocetin-induced platelet agglutination. To determine the capacity of activating platelets via the interaction with GPIbα, whole blood was incubated with the N-terminal region truncated or intact tri-A domain protein prior to perfusion over a fibrin(ogen)-coated surface. At a high shear rate of 1,500 s(-1), platelets from blood containing the truncated protein rapidly bound, covering >90% of the fibrin(ogen) surface area, whereas the intact tri-A domain protein induced platelets to bind <10%. The results obtained in this study ascertain the relevant role of the structural association between the N-terminal flanking region of the A1 domain (amino acids Gln(1238)-Glu(1260)) and the A1A2A3 domain complex in preventing vWF to bind spontaneously to GPIbα in solution under high shear forces.
authors.affiliation: Section of Cardiovascular Research, Department of Medicine, Baylor College of Medicine, Houston, Texas 77030, USA.
authors.forename: Matthew
authors.initials: M
authors.lastname: Auton
authors.forename: Katie E
authors.initials: KE
authors.lastname: Sowa
authors.forename: Molly
authors.initials: M
authors.lastname: Behymer
authors.forename: Miguel A
authors.initials: MA
authors.lastname: Cruz
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology; D005260:Female; D005340:Fibrinogen / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008297:Male; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism*; D046912:Multiprotein Complexes / Q000235:genetics / Q000378:metabolism; D015539:Platelet Activation / Q000187:drug effects / Q000502:physiology*; D019038:Platelet Glycoprotein GPIb-IX Complex; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D012310:Ristocetin / Q000494:pharmacology; D013312:Stress, Physiological / Q000187:drug effects / Q000502:physiology*; D014841:von Willebrand Factor / Q000235:genetics / Q000378:metabolism*
chemical_list: D000900:Anti-Bacterial Agents; D008562:Membrane Glycoproteins; D046912:Multiprotein Complexes; D019038:Platelet Glycoprotein GPIb-IX Complex; C094990:adhesion receptor; D014841:von Willebrand Factor; D012310:Ristocetin; D005340:Fibrinogen