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Blood. 2019 Apr 25;133(17):1909-1918. doi: 10.1182/blood-2018-11-886309. Epub 2019 Jan 28.

Exploring the "minimal" structure of a functional ADAMTS13 by mutagenesis and small-angle X-ray scattering.

Author information

1
Department of Medicine, Washington University School of Medicine, St. Louis, MO.
2
Laboratory for Thrombosis Research, Interdisciplinary Research Facility Life Sciences, Katholieke Universiteit Leuven Campus Kulak Kortrijk, Kortrijk, Belgium.
3
Laboratory of Malaria Immunology and Vaccinology, National Institute of Allergy and Infectious Disease, National Institutes of Health, Rockville, MD; and.
4
Department of Biochemistry and Molecular Biophysics, Washington University School of Medicine, St. Louis, MO.

Abstract

Human ADAMTS13 is a multidomain protein with metalloprotease (M), disintegrin-like (D), thrombospondin-1 (T), Cys-rich (C), and spacer (S) domains, followed by 7 additional T domains and 2 CUB (complement components C1r and C1s, sea urchin protein Uegf, and bone morphogenetic protein-1) domains. ADAMTS13 inhibits the growth of von Willebrand factor (VWF)-platelet aggregates by cleaving the cryptic Tyr1605-Met1606 bond in the VWF A2 domain. ADAMTS13 is regulated by substrate-induced allosteric activation; without shear stress, the distal T8-CUB domains markedly inhibit VWF cleavage, and binding of VWF domain D4 or selected monoclonal antibodies (MAbs) to distal ADAMTS13 domains relieves this autoinhibition. By small angle X-ray scattering (SAXS), ADAMTS13 adopts a hairpin-like conformation with distal T7-CUB domains close to the proximal MDTCS domains and a hinge point between T4 and T5. The hairpin projects like a handle away from the core MDTCS and T7-CUB complex and contains distal T domains that are dispensable for allosteric regulation. Truncated constructs that lack the T8-CUB domains are not autoinhibited and cannot be activated by VWF D4 but retain the hairpin fold. Allosteric activation by VWF D4 requires T7, T8, and the 58-amino acid residue linker between T8 and CUB1. Deletion of T3 to T6 produced the smallest construct (delT3-6) examined that could be activated by MAbs and VWF D4. Columba livia (pigeon) ADAMTS13 (pADAMTS13) resembles human delT3-6, retains normal activation by VWF D4, and has a SAXS envelope consistent with amputation of the hairpin containing the dispensable T domains of human ADAMTS13. Our findings suggest that human delT3-6 and pADAMTS13 approach a "minimal" structure for allosterically regulated ADAMTS13.

PMID:
30692120
PMCID:
PMC6484386
[Available on 2020-04-25]
DOI:
10.1182/blood-2018-11-886309

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