title: The functions of the A1A2A3 domains in von Willebrand factor include multimerin 1 binding.
abstract: Multimerin 1 (MMRN1) is a massive, homopolymeric protein that is stored in platelets and endothelial cells for activation-induced release. In vitro, MMRN1 binds to the outer surfaces of activated platelets and endothelial cells, the extracellular matrix (including collagen) and von Willebrand factor (VWF) to support platelet adhesive functions. VWF associates with MMRN1 at high shear, not static conditions, suggesting that shear exposes cryptic sites within VWF that support MMRN1 binding. Modified ELISA and surface plasmon resonance were used to study the structural features of VWF that support MMRN1 binding, and determine the affinities for VWF-MMRN1 binding. High shear microfluidic platelet adhesion assays determined the functional consequences for VWF-MMRN1 binding. VWF binding to MMRN1 was enhanced by shear exposure and ristocetin, and required VWF A1A2A3 region, specifically the A1 and A3 domains. VWF A1A2A3 bound to MMRN1 with a physiologically relevant binding affinity (KD: 2.0 ± 0.4 nM), whereas the individual VWF A1 (KD: 39.3 ± 7.7 nM) and A3 domains (KD: 229 ± 114 nM) bound to MMRN1 with lower affinities. VWF A1A2A3 was also sufficient to support the adhesion of resting platelets to MMRN1 at high shear, by a mechanism dependent on VWF-GPIbα binding. Our study provides new information on the molecular basis of MMRN1 binding to VWF, and its role in supporting platelet adhesion at high shear. We propose that at sites of vessel injury, MMRN1 that is released following activation of platelets and endothelial cells, binds to VWF A1A2A3 region to support platelet adhesion at arterial shear rates.
authors.forename: D'Andra N
authors.initials: DN
authors.lastname: Parker
authors.forename: Subia
authors.initials: S
authors.lastname: Tasneem
authors.forename: Richard W
authors.initials: RW
authors.lastname: Farndale
authors.forename: Dominique
authors.initials: D
authors.lastname: Bihan
authors.forename: J Evan
authors.initials: JE
authors.lastname: Sadler
authors.forename: Silvie
authors.initials: S
authors.lastname: Sebastian
authors.forename: Philip G
authors.initials: PG
authors.lastname: de Groot
authors.affiliation: Catherine P. M. Hayward, McMaster University Medical Centre, HSC 2N29A, 1200 Main St. West, Hamilton, Ontario, Canada L8N 3Z5, Tel.: +1 905 521 2100 Ext. 76274, Fax: +1 905 521 2338, E-mail: haywrdc@mcmaster.ca.
authors.forename: Catherine P M
authors.initials: CP
authors.lastname: Hayward
mesh_terms: D001792:Blood Platelets / Q000187:drug effects / Q000378:metabolism; D001798:Blood Proteins / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D066298:In Vitro Techniques; D046210:Microfluidic Analytical Techniques; D050505:Mutant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D015539:Platelet Activation; D010973:Platelet Adhesiveness; D019038:Platelet Glycoprotein GPIb-IX Complex / Q000037:antagonists & inhibitors / Q000378:metabolism; D011485:Protein Binding / Q000187:drug effects; D000072417:Protein Domains; D011994:Recombinant Proteins / Q000378:metabolism; D012310:Ristocetin / Q000494:pharmacology; D020349:Surface Plasmon Resonance; D014841:von Willebrand Factor / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
chemical_list: D001798:Blood Proteins; D050505:Mutant Proteins; D019038:Platelet Glycoprotein GPIb-IX Complex; D011994:Recombinant Proteins; C094990:adhesion receptor; C069282:multimerin; D014841:von Willebrand Factor; D012310:Ristocetin
keywords: adhesion molecules; collagens; platelet glycoproteins; von Willebrand factor