Independent assembly and secretion of a dimeric adhesive domain of von Willebrand factor containing the glycoprotein Ib-binding site.
von Willebrand factor (vWF) is a multimeric glycoprotein that supports platelet adhesion on thrombogenic surfaces as part of the normal hemostatic response to vascular injury. We have employed a domain-specific expression strategy to analyze the biosynthetic processing steps and minimum structural requirements for assembly of the platelet receptor glycoprotein Ib-binding domain of vWF. A chimeric cDNA that codes for the vWF signal peptide and a segment of vWF internal primary sequence, residues 441-730, directs the secretion of a functional vWF fragment from mammalian cells. The recombinant molecule intrinsically assembles through intermolecular disulfide bond formation into a dimeric adhesive domain without contributions from other regions of vWF, including propeptide, previously indicated as essential for vWF multimer assembly. Prevention of N-linked glycosylation on the recombinant domain does not impair dimer formation or the ability to support platelet aggregation. These results identify a minimum structural element for vWF subunit assembly and provide new insights into the processing steps to produce vWF multimers and adhesive domains.
Azuma
H
H
Room Research Laboratory for Arteriosclerosis and Thrombosis, Research Institute of Scripps Clinic, La Jolla, California 92037.
Dent
J A
JA
Sugimoto
M
M
Ruggeri
Z M
ZM
Ware
J
J
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002460:Cell Line; D002846:Chromatography, Affinity; D003094:Collagen / Q000037:antagonists & inhibitors; D006224:Cricetinae; D004247:DNA / Q000235:genetics; D006031:Glycosylation; D000882:Haplorhini; D006494:Heparin Antagonists / Q000378:metabolism; D008969:Molecular Sequence Data; D010957:Plasmids; D010974:Platelet Aggregation; D010980:Platelet Membrane Glycoproteins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D014170:Transformation, Genetic; D014415:Tunicamycin / Q000494:pharmacology; D014841:von Willebrand Factor / Q000235:genetics / Q000378:metabolism*
D006494:Heparin Antagonists; D010980:Platelet Membrane Glycoproteins; D014841:von Willebrand Factor; D014415:Tunicamycin; D003094:Collagen; D004247:DNA