title: Shear stress-induced unfolding of VWF accelerates oxidation of key methionine residues in the A1A2A3 region.
abstract: VWF is required for platelet adhesion to sites of vessel injury, a process vital for both hemostasis and thrombosis. Enhanced VWF secretion and oxidative stress are both hallmarks of inflammation. We recently showed that the neutrophil oxidant hypochlorous acid (HOCl) inhibits VWF proteolysis by ADAMTS13 by oxidizing VWF methionine 1606 (M1606) in the A2 domain. M1606 was readily oxidized in a substrate peptide, but required urea in multimeric plasma VWF. In the present study, we examined whether shear stress enhances VWF oxidation. With an HOCl-generating system containing myeloperoxidase (MPO) and H(2)O(2), we found that shear stress accelerated M1606 oxidation, with 56% becoming oxidized within 1 hour. Seven other methionine residues in the VWF A1A2A3 region (containing the sites for platelet and collagen binding and ADAMTS13 cleavage) were variably oxidized, one completely. Oxidized methionines accumulated preferentially in the largest VWF multimers. HOCl-oxidized VWF was hyperfunctional, agglutinating platelets at ristocetin concentrations that induced minimal agglutination using unoxidized VWF and binding more of the nanobody AU/VWFa-11, which detects a gain-of-function conformation of the A1 domain. These findings suggest that neutrophil oxidants will both render newly secreted VWF uncleavable and alter the largest plasma VWF forms such that they become hyperfunctional and resistant to proteolysis by ADAMTS13.
authors.affiliation: Research Division, Puget Sound Blood Center, Seattle, WA 98104, USA. josel@psbcresearch.org
authors.forename: Xiaoyun
authors.initials: X
authors.lastname: Fu
authors.forename: Junmei
authors.initials: J
authors.lastname: Chen
authors.forename: Ryan
authors.initials: R
authors.lastname: Gallagher
authors.forename: Ying
authors.initials: Y
authors.lastname: Zheng
authors.forename: Dominic W
authors.initials: DW
authors.lastname: Chung
authors.forename: José A
authors.initials: JA
authors.lastname: López
mesh_terms: D051722:ADAM Proteins / Q000378:metabolism; D000071120:ADAMTS13 Protein; D001665:Binding Sites; D001792:Blood Platelets / Q000378:metabolism; D006801:Humans; D006997:Hypochlorous Acid; D066298:In Vitro Techniques; D008715:Methionine / Q000737:chemistry; D008958:Models, Molecular; D010084:Oxidation-Reduction; D055503:Protein Multimerization; D058767:Protein Unfolding; D059748:Proteolysis; D033081:Shear Strength; D013314:Stress, Mechanical; D014841:von Willebrand Factor / Q000737:chemistry* / Q000378:metabolism
chemical_list: D014841:von Willebrand Factor; D006997:Hypochlorous Acid; D008715:Methionine; D051722:ADAM Proteins; D000071120:ADAMTS13 Protein; C099604:ADAMTS13 protein, human