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Crystal structures of the noncatalytic domains of ADAMTS13 reveal multiple discontinuous exosites for von Willebrand factor.

ABSTRACT: ADAMTS13 specifically cleaves plasma von Willebrand factor (VWF) and thereby controls VWF-mediated platelet thrombus formation. Severe deficiencies in ADAMTS13 can cause life-threatening thrombotic thrombocytopenic purpura. Here, we determined 2 crystal structures of ADAMTS13-DTCS (residues 287-685), an exosite-containing human ADAMTS13 fragment, at 2.6-A and 2.8-A resolution. The structures revealed folding similarities between the disintegrin-like (D) domain and the N-terminal portion of the cysteine-rich domain (designated the C(A) domain). The spacer (S) domain forms a globular functional unit with a 10-stranded beta-sandwich fold that has multiple interaction sites with the C(A) domain. We expressed 25 structure-based mutants of ADAMTS13-MDTCS (residues 75-685) and measured their enzymatic activity. We identified 3 VWF-binding exosites on the linearly aligned discontinuous surfaces of the D, C(A), and S domains traversing the W-shaped molecule. Since the MDTCS domains are conserved among ADAMTS family proteins, the structural framework of the multiple enzyme-substrate interactions identified in the ADAMTS13-VWF system provides the basis for a common substrate recognition mode in this class of proteinases.

SUBMITTER: Akiyama M

PROVIDER: S-EPMC2780749 | biostudies-literature | 2009 Nov

REPOSITORIES: biostudies-literature

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Crystal structures of the noncatalytic domains of ADAMTS13 reveal multiple discontinuous exosites for von Willebrand factor.

Akiyama Masashi M Takeda Soichi S Kokame Koichi K Takagi Junichi J Miyata Toshiyuki T

Proceedings of the National Academy of Sciences of the United States of America 20091030 46

ADAMTS13 specifically cleaves plasma von Willebrand factor (VWF) and thereby controls VWF-mediated platelet thrombus formation. Severe deficiencies in ADAMTS13 can cause life-threatening thrombotic thrombocytopenic purpura. Here, we determined 2 crystal structures of ADAMTS13-DTCS (residues 287-685), an exosite-containing human ADAMTS13 fragment, at 2.6-A and 2.8-A resolution. The structures revealed folding similarities between the disintegrin-like (D) domain and the N-terminal portion of the c ...[more]

PMID: 19880749

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Project description:Essentials The mechanism for the auto-inhibition of von Willebrand factor (VWF) remains unclear. Hydrogen exchange of two VWF A1 fragments with disparate activities was measured and compared. Discontinuous residues flanking A1 form a structural module that blocks A1 binding to the platelet. Our results suggest a potentially unified model of VWF activation. Click to hear an ISTH Academy presentation on the domain architecture of VWF and activation by elongational flow by Dr Springer SUMMARY: Background How von Willebrand factor (VWF) senses and responds to shear flow remains unclear. In the absence of shear flow, VWF or its fragments can be induced to bind spontaneously to platelet GPIb?. Objectives To elucidate the auto-inhibition mechanism of VWF. Methods Hydrogen-deuterium exchange (HDX) of two recombinant VWF fragments expressed from baby hamster kidney cells were measured and compared. Results The shortA1 protein contains VWF residues 1261-1472 and binds GPIb? with a significantly higher affinity than the longA1 protein that contains VWF residues 1238-1472. Both proteins contain the VWF A1 domain (residues 1272-1458). Many residues in longA1, particularly those in the N- and C-terminal sequences flanking the A1 domain, and in helix ?1, loops ?1?2 and ?3?2, demonstrated markedly reduced HDX compared with their counterparts in shortA1. The HDX-protected region in longA1 overlaps with the GPIb?-binding interface and is clustered with type 2B von Willebrand disease (VWD) mutations. Additional comparison with the HDX of denatured longA1 and ristocetin-bound longA1 indicates the N- and C-terminal sequences flanking the A1 domain form cooperatively an integrated autoinhibitory module (AIM) that interacts with the HDX-protected region. Binding of ristocetin to the C-terminal part of the AIM desorbs the AIM from A1 and enables longA1 binding to GPIb?. Conclusion The discontinuous AIM binds the A1 domain and prevents it from binding to GPIb?, which has significant implications for the pathogenesis of type 2B VWD and the shear-induced activation of VWF activity.

Dataset Information

Crystal structures of the noncatalytic domains of ADAMTS13 reveal multiple discontinuous exosites for von Willebrand factor.

Publications

Crystal structures of the noncatalytic domains of ADAMTS13 reveal multiple discontinuous exosites for von Willebrand factor.

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