title: A novel binding site for ADAMTS13 constitutively exposed on the surface of globular VWF.
abstract: ADAMTS13 metalloprotease regulates the multimeric size of von Willebrand factor (VWF) by cleaving the Tyr1605-Met1606 bond in the VWF A2 domain. The mechanisms of VWF recognition by ADAMTS13 have yet to be fully resolved. Most studies have focused on the role of exosites within the VWF A2 domain, involved in interaction with the ADAMTS13 spacer domain. In the present study, we expressed different C-terminal domain VWF fragments and evaluated their binding to ADAMTS13 and its truncated mutants, MDTCS and del(TSP5-CUB). Using plate binding assay and surface plasmon resonance, we identified a novel ADAMTS13 binding site (K(D) approximately 86 nM) in the region of VWF spanning residues 1874 to 2813, which includes the VWF D4 domain and that interacts with the C-terminal domains of ADAMTS13. We show that the interaction occurs even when VWF is in static conditions, assumed to be globular and where the VWF A2 domain is hidden. We demonstrate that C-terminal VWF fragments, as well as an antibody specifically directed toward the VWF D4 domain, inhibit VWF proteolysis by ADAMTS13 under shear conditions. We propose that this novel VWF C-terminal binding site may participate as the initial step of a multistep interaction ultimately leading to proteolysis of VWF by ADAMTS13.
authors: Department of Haematology, Imperial College London, Hammersmith Hospital Campus, London, United Kingdom. sara.zanardelli03@imperial.ac.uk
authors: Sara
authors: S
authors: Zanardelli
authors: Alain C K
authors: AC
authors: Chion
authors: Evelyn
authors: E
authors: Groot
authors: Peter J
authors: PJ
authors: Lenting
authors: Thomas A J
authors: TA
authors: McKinnon
authors: Mike A
authors: MA
authors: Laffan
authors: Michelle
authors: M
authors: Tseng
authors: David A
authors: DA
authors: Lane
mesh_terms: D051722:ADAM Proteins / Q000737:chemistry / Q000378:metabolism*; D000071120:ADAMTS13 Protein; D001665:Binding Sites; D002478:Cells, Cultured; D006801:Humans; D008954:Models, Biological; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D017510:Protein Folding; D025941:Protein Interaction Mapping; D011499:Protein Processing, Post-Translational; D017434:Protein Structure, Tertiary / Q000502:physiology; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D014841:von Willebrand Factor / Q000737:chemistry* / Q000378:metabolism*
chemical_list: D010446:Peptide Fragments; D011994:Recombinant Proteins; D014841:von Willebrand Factor; D051722:ADAM Proteins; D000071120:ADAMTS13 Protein; C099604:ADAMTS13 protein, human