Formation of methionine sulfoxide by peroxynitrite at position 1606 of von Willebrand factor inhibits its cleavage by ADAMTS-13: A new prothrombotic mechanism in diseases associated with oxidative stress |
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| Authors: | Stefano Lancellotti Vincenzo De Filippis Nicola Pozzi Flora Peyvandi Roberta Palla Bianca Rocca Sergio Rutella Dario Pitocco Pier Mannuccio Mannucci Raimondo De Cristofaro |
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| Institution: | 1. Division of Epidemiology and Community Health, School of Public Health, University of Minnesota, Minneapolis, MN, United States;2. Lipid and Atherosclerosis Unit, IMIBIC/Hospital Universitario Reina Sofía/Universidad de Córdoba, Córdoba, Spain;3. CIBER Fisiopatología Obesidad y Nutrición (CIBEROBN), Instituto de Salud Carlos III, Madrid, Spain;4. Cardiovascular Division, Department of Medicine, University of Minnesota Medical School, Minneapolis, MN, United States;5. Wake Forest University, Winston-Salem, NC, United States;6. Department of Medicine, Baylor College of Medicine, Houston, TX, United States;7. McKusick-Nathans Institute of Genetic Medicine, Johns Hopkins University School of Medicine, Baltimore, MD, United States;8. Department of Epidemiology, Gillings School of Global Public Health, University of North Carolina, Chapel Hill, NC, United States;9. Department of Epidemiology, Rollins School of Public Health, Emory University, Atlanta, GA, United States;2. Department of Hematology, First Medical Center, General Hospital of the PLA, Beijing, China;3. Department of Orthopedics, First Medical Center, General Hospital of the PLA, Beijing, China;4. Department of Ultrasound, Hainan Hospital of the PLA General Hospital, Sanya, China;1. University Children''s Hospital Zurich, Switzerland;2. Neuroscience Center Zurich (ZNZ), Switzerland;3. Zurich Center for Integrative Human Physiology (ZIHP), University of Zurich, Switzerland;4. Pulmonary Division, University Hospital Zurich, Switzerland;5. Institute of Pharmacology and Toxicology, University of Zurich, Switzerland;6. University Clinics for Child and Adolescent Psychiatry, Zurich, Switzerland |
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| Abstract: | An enhanced formation of reactive oxygen species and peroxynitrite occurs in several clinical settings including diabetes, coronary artery disease, stroke, sepsis, and chronic inflammatory diseases. Peroxynitrite oxidizes methionine and tyrosine residues to methionine sulfoxide (MetSO) and 3-nitrotyrosine (NT), respectively. Notably, ADAMTS-13 cleaves von Willebrand factor (VWF) exclusively at the Tyr1605–Met1606 peptide bond in the A2 domain. We hypothesized that peroxynitrite could oxidize either or both of these amino acid residues, thus potentially affecting ADAMTS-13-mediated cleavage. We tested our hypothesis using synthetic peptide substrates based on: (1) VWF Asp1596–Ala1669 sequence (VWF74) and (2) VWF Asp1596–Ala1669 sequence containing nitrotyrosine (VWF74-NT) or methionine sulfoxide (VWF74-MetSO) at position 1605 or 1606, respectively. The peptides were treated with recombinant ADAMTS-13 and the cleavage products analyzed by RP-HPLC. VWF74 oxidized by peroxynitrite underwent a severe impairment of its hydrolysis. Likewise, VWF74-MetSO was minimally hydrolyzed, whereas VWF74-NT was hydrolyzed slightly more efficiently than VWF74. Oxidation by peroxynitrite of purified VWF multimers inhibited ADAMTS-13 hydrolysis, but did not alter their electrophoretic pattern nor their ability to induce platelet agglutination by ristocetin. Moreover, VWF purified from type 2 diabetic patients showed oxidative damage, as revealed by enhanced carbonyl, NT, and MetSO content and was partially resistant to ADAMTS-13 hydrolysis. In conclusion, peroxynitrite may contribute to prothrombotic effects, hindering the proteolytic processing by ADAMTS-13 of high-molecular-weight VWF multimers, which have the highest ability to bind and activate platelets in the microcirculation. |
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