title: Shear-induced unfolding activates von Willebrand factor A2 domain for proteolysis.
abstract: BACKGROUND
  To avoid pathological platelet aggregation by von Willebrand factor (VWF), VWF multimers are regulated in size and reactivity for adhesion by ADAMTS13-mediated proteolysis in a shear flow dependent manner.
  
  
  OBJECTIVE
  We examined whether tensile stress in VWF under shear flow activates the VWF A2 domain for cleavage by ADAMTS13 using molecular dynamics simulations. We generated a full length mutant VWF featuring a homologous disulfide bond in A2 (N1493C and C1670S), in an attempt to lock A2 against unfolding.
  
  
  RESULTS
  We indeed observed stepwise unfolding of A2 and exposure of its deeply buried ADAMTS13 cleavage site. Interestingly, disulfide bonds in the adjacent and highly homologous VWF A1 and A3 domains obstruct their mechanical unfolding. We find this mutant A2 (N1493C and C1670S) to feature ADAMTS13-resistant behavior in vitro.
  
  
  CONCLUSIONS
  Our results yield molecular-detail evidence for the force-sensing function of VWF A2, by revealing how tension in VWF due to shear flow selectively exposes the A2 proteolysis site to ADAMTS13 for cleavage while keeping the folded remainder of A2 intact and functional. We find the unconventional 'knotted' Rossmann fold of A2 to be the key to this mechanical response, tailored for regulating VWF size and activity. Based on our model we discuss the pathomechanism of some natural mutations in the VWF A2 domain that significantly increase the cleavage by ADAMTS13 without shearing or chemical denaturation, and provide with the cleavage-activated A2 conformation a structural basis for the design of inhibitors for VWF type 2 diseases.
authors.affiliation: CAS-MPG Partner Institute for Computational Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, China. carsten@picb.ac.cn
authors.forename: C
authors.initials: C
authors.lastname: Baldauf
authors.forename: R
authors.initials: R
authors.lastname: Schneppenheim
authors.forename: W
authors.initials: W
authors.lastname: Stacklies
authors.forename: T
authors.initials: T
authors.lastname: Obser
authors.forename: A
authors.initials: A
authors.lastname: Pieconka
authors.forename: S
authors.initials: S
authors.lastname: Schneppenheim
authors.forename: U
authors.initials: U
authors.lastname: Budde
authors.forename: J
authors.initials: J
authors.lastname: Zhou
authors.forename: F
authors.initials: F
authors.lastname: Gräter
mesh_terms: D051722:ADAM Proteins / Q000378:metabolism*; D000071120:ADAMTS13 Protein; D000268:Adhesiveness; D006801:Humans; D006868:Hydrolysis; D009154:Mutation; D011485:Protein Binding; D011487:Protein Conformation; D011489:Protein Denaturation; D055503:Protein Multimerization; D055550:Protein Stability; D013314:Stress, Mechanical; D014841:von Willebrand Factor / Q000235:genetics / Q000378:metabolism*
chemical_list: D014841:von Willebrand Factor; D051722:ADAM Proteins; D000071120:ADAMTS13 Protein; C099604:ADAMTS13 protein, human