Etude des relations structure-fonctions du couple facteur Willebrand-ADAMTS13

Von Willebrand factor is a multimeric glycoprotein essential for platelet adhesion and aggregation. Multimerization degree is regulated in vivo par ADAMTS13. Deficiency in ADAMTS13 activity induces the thrombotic thrombocytopenic purpura (TTP). Deficiency in VWF induces the Von Willebrand disease (VWD). The aim of this study is to establish the role of ADAMTS13 in the physiopathology of VWD type 2 and hereditary TTP and to specify VWF domains implicated in the recognition of ADAMTS13. Sensitivity of recombinant (r) VWF type 2 to proteolysis by rADAMTS13 was investigated. Using low ionic strength conditions, all mutations induced increased proteolysis of rVWF by ADAMTS13. At physiological salt concentration, type 2A and type 2B mutants exhibited a significantly higher susceptibility to ADAMTS13, whereas sensitivity of type 2M mutants to ADAMTS13 was normalized. In VWD type 2B, the spontaneous binding to platelets and excessive degradation by ADAMTS13 of VWF high-molecular-weight multimers may account for their clearance from plasma. Using a panel of VWF-fragments overlapping the entire VWF subunit and deleted-VWF, two distinct regions in von Willebrand factor contribute to binding of ADAMTS13: A major binding site within the A2 domain and a primary but auxiliary binding site within the A3 domain. The second part of this thesis report the study of 4 mutations of ADAMTS13 identified in hereditary TPP patients. We established that these mutations were responsible for the disease. The defect of ADAMTS13 activity is even qualitative, associated with reduced binding to VWF, even quantitative,resulting of intracellular retention.