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A model for the conformational activation of the structurally quiescent metalloprotease ADAMTS13 by Von Willebrand factor

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Title: A model for the conformational activation of the structurally quiescent metalloprotease ADAMTS13 by Von Willebrand factor
Authors: South, K
Freitas, MO
Lane, DA
Item Type: Journal Article
Abstract: Blood loss is prevented by the multi-domain glycoprotein von Willebrand factor (VWF), which binds exposed collagen at damaged vessels and captures platelets. VWF is regulated by the metalloprotease ADAMTS13, which, in turn, is conformationally activated by VWF. To delineate the structural requirements for VWF-mediated conformational activation of ADAMTS13, we performed binding and functional studies with a panel of truncated ADAMTS13 variants. We demonstrate that both the isolated CUB1 and CUB2 domains in ADAMTS13 bind to the spacer domain exosite of a truncated ADAMTS13 variant, MDTCS (KD of 135 ± 10.1 nM and 86.9 ± 9.0 nM, respectively). However, only the CUB1 domain inhibited proteolytic activity of MDTCS. Moreover, ADAMTS13∆CUB2, unlike ADAMTS13∆CUB1-2, exhibited activity similar to wild-type ADAMTS13 and could be activated by VWF D4-CK. The CUB2 domain is therefore not essential for maintaining the inactive conformation of ADAMTS13. Both CUB domains could bind to the VWF D4-CK domain fragment (KD of 53.7± 2.1 nM and 84.3 ± 2.0 nM, respectively). However, deletion of both CUB domains did not prevent VWF D4-CK binding, suggesting that competition for CUB-domain binding to the spacer domain is not the dominant mechanism behind the conformational activation. ADAMTS13∆TSP8-CUB2 could no longer bind to VWF D4-CK, and deletion of TSP8 abrogated ADAMTS13 conformational activation. These findings support an ADAMTS13-activation model in which VWF D4-CK engages the TSP8-CUB2 domains, inducing the conformational change that disrupts the CUB1-spacer domain interaction and thereby activates ADAMTS13.
Issue Date: 16-Feb-2017
Date of Acceptance: 16-Feb-2017
URI: http://hdl.handle.net/10044/1/44757
DOI: https://dx.doi.org/10.1074/jbc.M117.776732
ISSN: 0021-9258
Publisher: American Society for Biochemistry and Molecular Biology
Start Page: 5760
End Page: 5769
Journal / Book Title: Journal of Biological Chemistry
Volume: 292
Issue: 14
Copyright Statement: © 2017 by The American Society for Biochemistry and Molecular Biology, Inc. Final version free via Creative Commons CC-BY license.
Sponsor/Funder: British Heart Foundation
British Heart Foundation
Funder's Grant Number: PG/12/55/29740
PG/14/87/3118
Keywords: Science & Technology
Life Sciences & Biomedicine
Biochemistry & Molecular Biology
ADAMTS13
allosteric regulation
hemostasis
protein conformation
von Willebrand factor
THROMBOTIC THROMBOCYTOPENIC PURPURA
PLATELET GLYCOPROTEIN-IB
FACTOR A2 DOMAIN
VONWILLEBRAND-FACTOR
ISCHEMIC-STROKE
SCISSILE BOND
MYOCARDIAL-INFARCTION
VICINAL CYSTEINES
BINDING-SITES
HUMAN-PLASMA
ADAMTS13 Protein
HEK293 Cells
Humans
Models, Chemical
Protein Binding
Protein Domains
von Willebrand Factor
06 Biological Sciences
11 Medical And Health Sciences
03 Chemical Sciences
Publication Status: Published
Appears in Collections:Department of Medicine
Faculty of Medicine



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