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Antibodies that conformationally activate ADAMTS13 allosterically enhance metalloprotease domain function

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Title: Antibodies that conformationally activate ADAMTS13 allosterically enhance metalloprotease domain function
Authors: Schelpe, A-S
Petri, A
Roose, E
Pareyn, I
Deckmyn, H
De Meyer, SF
Crawley, JTB
Vanhoorelbeke, K
Item Type: Journal Article
Abstract: Plasma ADAMTS13 circulates in a folded conformation that is stabilized by an interaction between the central Spacer domain and the C-terminal CUB (complement components C1r and C1s, sea urchin protein Uegf, and bone morphogenetic protein-1) domains. Binding of ADAMTS13 to the VWF D4(-CK) domains or to certain activating murine monoclonal antibodies (mAbs) induces a structural change that extends ADAMTS13 into an open conformation that enhances its function. The objective was to characterize the mechanism by which conformational activation enhances ADAMTS13-mediated proteolysis of VWF. The activating effects of a novel anti-Spacer (3E4) and the anti-CUB1 (17G2) mAbs on the kinetics of proteolysis of VWF A2 domain fragments by ADAMTS13 were analyzed. mAb-induced conformational changes in ADAMTS13 were investigated by enzyme-linked immunosorbent assay. Both mAbs enhanced ADAMTS13 catalytic efficiency (kcat/Km) by ∼twofold (3E4: 2.0-fold; 17G2: 1.8-fold). Contrary to previous hypotheses, ADAMTS13 activation was not mediated through exposure of the Spacer or cysteine-rich domain exosites. Kinetic analyses revealed that mAb-induced conformational extension of ADAMTS13 enhances the proteolytic function of the metalloprotease domain (kcat), rather than augmenting substrate binding (Km). A conformational effect on the metalloprotease domain was further corroborated by the finding that incubation of ADAMTS13 with either mAb exposed a cryptic epitope in the metalloprotease domain that is normally concealed when ADAMTS13 is in a closed conformation. We show for the first time that the primary mechanism of mAb-induced conformational activation of ADAMTS13 is not a consequence of functional exosite exposure. Rather, our data are consistent with an allosteric activation mechanism on the metalloprotease domain that augments active site function.
Issue Date: 24-Mar-2020
Date of Acceptance: 11-Feb-2020
URI: http://hdl.handle.net/10044/1/77752
DOI: 10.1182/bloodadvances.2019001375
ISSN: 2473-9529
Publisher: American Society of Hematology
Start Page: 1072
End Page: 1080
Journal / Book Title: Blood Advances
Volume: 4
Issue: 6
Copyright Statement: © 2020 by The American Society of Hematology
Sponsor/Funder: British Heart Foundation
Funder's Grant Number: PG/18/17/33572
Publication Status: Published
Online Publication Date: 2020-03-20
Appears in Collections:Department of Immunology and Inflammation