title: Lateral self-association of VWF involves the Cys2431-Cys2453 disulfide/dithiol in the C2 domain.
abstract: VWF is a plasma protein that binds platelets to an injured vascular wall during thrombosis. When exposed to the shear forces found in flowing blood, VWF molecules undergo lateral self-association that results in a meshwork of VWF fibers. Fiber formation has been shown to involve thiol/disulfide exchange between VWF molecules. A C-terminal fragment of VWF was expressed in mammalian cells and examined for unpaired cysteine thiols using tandem mass spectrometry (MS). The VWF C2 domain Cys2431-Cys2453 disulfide bond was shown to be reduced in approximately 75% of the molecules. Fragments containing all 3 C domains or just the C2 domain formed monomers, dimers, and higher-order oligomers when expressed in mammalian cells. Mutagenesis studies showed that both the Cys2431-Cys2453 and nearby Cys2451-Cys2468 disulfide bonds were involved in oligomer formation. Our present findings imply that lateral VWF dimers form when a Cys2431 thiolate anion attacks the Cys2431 sulfur atom of the Cys2431-Cys2453 disulfide bond of another VWF molecule, whereas the Cys2451-Cys2468 disulfide/dithiol mediates formation of trimers and higher-order oligomers. These observations provide the basis for exploring defects in lateral VWF association in patients with unexplained hemorrhage or thrombosis.
authors.affiliation: Adult Cancer Program, Lowy Cancer Research Centre and Prince of Wales Clinical School, University of New South Wales, Sydney, Australia.
authors.forename: Tim
authors.initials: T
authors.lastname: Ganderton
authors.forename: Jason W H
authors.initials: JW
authors.lastname: Wong
authors.forename: Christina
authors.initials: C
authors.lastname: Schroeder
authors.forename: Philip J
authors.initials: PJ
authors.lastname: Hogg
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D003553:Cystine / Q000737:chemistry; D019281:Dimerization; D004220:Disulfides / Q000737:chemistry; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics; D055503:Protein Multimerization; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics; D014841:von Willebrand Factor / Q000737:chemistry* / Q000235:genetics
chemical_list: D004220:Disulfides; D010446:Peptide Fragments; D011994:Recombinant Proteins; D014841:von Willebrand Factor; D003553:Cystine