Phylogenetic and functional analysis of ADAMTS13 identifies highly conserved domains essential for allosteric regulation

Joshua Muia, Jian Zhu, Suellen C. Greco, Karen Vanhoorelbeke, Garima Gupta, Lisa A. Westfield and J. Evan Sadler

Key points

  • ADAMTS13 structure is exceptionally variable among vertebrates.

  • Functional, fully allosteric regulated ADAMTS13 requires no more than 3 distal thrombospondin repeats.


The metalloprotease ADAMTS13 prevents microvascular thrombosis by cleaving von Willebrand factor (VWF) within platelet-rich thrombi, and cleavage depends on allosteric activation of ADAMTS13 by the substrate VWF. Human ADAMTS13 has a short propeptide, metalloprotease (M), disintegrin-like (D), thrombospondin-1 (T), Cys-rich (C) and spacer (S) domains (proximal domains), followed by 7 T and 2 CUB domains (distal domains). Distal domains inhibit the catalytic proximal domains; binding of distal T8-CUB domains to the VWF D4 domain relieves autoinhibition and promotes cleavage of the nearby VWF A2 domain. However, the role of specific ADAMTS13 distal domains in this allosteric mechanism is not established. Assays of plasma ADAMTS13 from 20 placental mammals, birds and amphibians show that allosteric regulation is broadly conserved, and phylogenetic analysis of 264 vertebrates shows the long propeptide, T3, T4, T6 and T6a domains have been deleted several times in placental mammals, birds, and fish. Notably, pigeon ADAMTS13 has only 3 distal T domains but was activated normally by human VWF D4 and cleaved VWF multimers preferentially under fluid shear stress. Human ADAMTS13 constructed to resemble pigeon ADAMTS13 retained normal allosteric regulation and shear-dependent cleavage of VWF. Thus, the T3-T6 domains of human ADAMTS13 are dispensable. Conversely, deletion of T7 or T8 abolished allosteric activation. For most species, some sequence changes in the VWF substrate can markedly increase the rate of cleavage, suggesting that ADAMTS13 and VWF have not evolved to be optimal enzyme-substrate pairs. These properties may reflect evolutionary pressure to balance the risk of VWF-dependent bleeding and thrombosis.

  • Submitted November 15, 2018.
  • Accepted January 20, 2019.