The importance of vicinal cysteines, C1669 and C1670, for von Willebrand factor A2 domain function

Abstract The von Willebrand factor (VWF) A2 crystal structure has revealed the presence of a rare vicinal disulfide bond between C1669 and C1670, predicted to influence domain unfolding required for proteolysis by ADAMTS13. We prepared VWF A2 domain fragments with (A2-VicCC, residues 1473-1670) and without the vicinal disulfide bond (A2-ΔCC, residues 1473-1668). Compared with A2-ΔCC, A2-VicCC exhibited impaired proteolysis and also reduced binding to ADAMTS13. Circular dichroism studies revealed that A2-VicCC was more resistant to thermal unfolding than A2-ΔCC. Mutagenesis of C1669/C1670 in full-length VWF resulted in markedly increased susceptibility to cleavage by ADAMTS13, confirming the important role of the paired vicinal cysteines in VWF A2 domain stabilization.

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Expression of a structurally constrained von Willebrand factor variant triggers acute thrombotic thrombocytopenic purpura in mice

Blood ◽

10.1182/blood-2013-10-531392 ◽

2014 ◽

Vol 123 (21) ◽

pp. 3344-3353 ◽

Cited By ~ 6

Author(s):

Yoko Morioka ◽

Caterina Casari ◽

Nikolett Wohner ◽

Sungyun Cho ◽

Sachiko Kurata ◽

...

Keyword(s):

Thrombotic Thrombocytopenic Purpura ◽

Disulfide Bond ◽

Von Willebrand Factor ◽

Acute Onset ◽

Thrombocytopenic Purpura ◽

Key Points ◽

Von Willebrand ◽

A2 Domain ◽

Willebrand Factor

Key Points Introduction of a disulfide bond within the A2 domain renders VWF highly thrombogenic and resistant to proteolysis. Expression of mVWF/p.S1494C-p.A1534C in mice triggers an acute onset of thrombotic thrombocytopenic purpura.

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The disulfide bond Cys2724-Cys2774 in the C-terminal cystine knot domain of von Willebrand factor is critical for its dimerization and secretion

Thrombosis Journal ◽

10.1186/s12959-021-00348-w ◽

2021 ◽

Vol 19 (1) ◽

Author(s):

Yuxin Zhang ◽

Fengwu Chen ◽

Aizhen Yang ◽

Xiaoying Wang ◽

Yue Han ◽

...

Keyword(s):

Disulfide Bond ◽

Von Willebrand Factor ◽

Disulfide Bonds ◽

Von Willebrand Disease ◽

Cystine Knot ◽

Type 3 ◽

Von Willebrand ◽

Willebrand Factor

Abstract Background Type 3 von Willebrand disease (VWD) exhibits severe hemorrhagic tendency with complicated pathogenesis. The C-terminal cystine knot (CTCK) domain plays an important role in the dimerization and secretion of von Willebrand factor (VWF). The CTCK domain has four intrachain disulfide bonds including Cys2724-Cys2774, Cys2739-Cys2788, Cys2750-Cys2804 and Cys2754-Cys2806, and the single cysteine mutation in Cys2739-Cys2788, Cys2750-Cys2804 and Cys2754-Cys2806 result in type 3 VWD, demonstrating the crucial role of these three disulfide bonds in VWF biosynthesis, however, the role of the remaining disulfide bond Cys2724-Cys2774 remains unclear. Method and results In this study, by the next-generation sequencing we found a missense mutation a c.8171G>A (C2724Y) in the CTCK domain of VWF allele in a patient family with type 3 VWD. In vitro, VWF C2724Y protein was expressed normally in HEK-293T cells but did not form a dimer or secrete into cell culture medium, suggesting that C2724 is critical for the VWF dimerization, and thus for VWF multimerization and secretion. Conclusions Our findings provide the first genetic evidence for the important role of Cys2724-Cys2774 in VWF biosynthesis and secretion. Therefore, all of the four intrachain disulfide bonds in CTCK monomer contribute to VWF dimerization and secretion.

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Role of calcium in regulating the intra- and extracellular cleavage of von Willebrand factor by the protease ADAMTS13

Blood Advances ◽

10.1182/bloodadvances.2017009027 ◽

2017 ◽

Vol 1 (23) ◽

pp. 2063-2074 ◽

Cited By ~ 8

Author(s):

Shobhit Gogia ◽

Anju Kelkar ◽

Changjie Zhang ◽

Kannayakanahalli M. Dayananda ◽

Sriram Neelamegham

Keyword(s):

Von Willebrand Factor ◽

Calcium Binding ◽

Key Points ◽

Platelet Binding ◽

Von Willebrand ◽

A2 Domain ◽

Willebrand Factor ◽

Intracellular Proteolysis

Key Points VWF A2-domain intracellular proteolysis within ECs is enhanced upon disrupting calcium binding (eg, in VWD type 2A mutants). VWF string cleavage on ECs is calcium independent and is strongly dependent on platelet binding.

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Role of Platelets, Thrombin, Integrin llb-llla, Fibronectin and Von Willebrand Factor on Tumor Adhesion in Vitro and Metastasis in Vivo

Thrombosis and Haemostasis ◽

10.1055/s-0038-1642691 ◽

1995 ◽

Vol 74 (01) ◽

pp. 282-290 ◽

Cited By ~ 99

Author(s):

Mary Lynn Nierodzik ◽

Abraham Klepfish ◽

Simon Karpatkin

Keyword(s):

Von Willebrand Factor ◽

Von Willebrand ◽

Tumor Adhesion ◽

Willebrand Factor

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Factor VIII Inhibitor Antibodies with C2 Domain Specificity Are Less inhibitory to Factor VIII Complexed with von Willebrand Factor

Thrombosis and Haemostasis ◽

10.1055/s-0038-1650655 ◽

1996 ◽

Vol 76 (05) ◽

pp. 749-754 ◽

Cited By ~ 73

Author(s):

Suzuki Suzuki ◽

Morio Arai ◽

Kagehiro Amano ◽

Kazuhiko Kagawa ◽

Katsuyuki Fukutake

Keyword(s):

Complex Formation ◽

Factor Viii ◽

Von Willebrand Factor ◽

Domain Specificity ◽

Factor Viii Inhibitor ◽

C2 Domain ◽

Recombinant Fviii ◽

Von Willebrand ◽

A2 Domain ◽

Willebrand Factor

SummaryIn order to clarify the potential role of von Willebrand factor (vWf) in attenuating the inactivation of factor VIII (fVIII) by those antibodies with C2 domain specificity, we investigated a panel of 14 human antibodies to fVIII. Immunoblotting analysis localized light chain (C2 domain) epitopes for four cases, heavy chain (A2 domain) epitopes in five cases, while the remaining five cases were both light and heavy chains. The inhibitor titer was considerably higher for Kogenate, a recombinant fVIII concentrate, than for Haemate P, a fVIII/vWf complex concentrate, in all inhibitor plasmas that had C2 domain specificity. In five inhibitor plasmas with A2 domain specificity and in five with both A2 and C2 domain specificities, Kogenate gave titers similar to or lower than those with Haemate P. The inhibitory effect of IgG of each inhibitor plasma was then compared with recombinant fVIII and its complex with vWf. When compared to the other 10 inhibitor IgGs, IgG concentration, which inhibited 50% of fVIII activity (IC50), was remarkably higher for the fVIII/vWf complex than for fVIII in all the inhibitor IgGs that had C2 domain reactivity. Competition of inhibitor IgG and vWf for fVIII binding was observed in an ELISA system. In 10 inhibitors that had C2 domain reactivity, the dose dependent inhibition of fVIII-vWf complex formation was observed, while, in the group of inhibitors with A2 domain specificity, there was no inhibition of the complex formation except one case. We conclude that a subset of fVIII inhibitors, those that bind to C2 domain determinants, are less inhibitory to fVIII when it is complexed with vWf that binds to overlapping region in the C2 domain.

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Faculty Opinions recommendation of Alloantibodies to therapeutic factor VIII in hemophilia A: the role of von Willebrand factor in regulating factor VIII immunogenicity.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.725335385.793503724 ◽

2015 ◽

Author(s):

Gili Kenet

Keyword(s):

Factor Viii ◽

Von Willebrand Factor ◽

Hemophilia A ◽

Von Willebrand ◽

Willebrand Factor

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The role of von Willebrand factor in breast cancer metastasis

Translational Oncology ◽

10.1016/j.tranon.2021.101033 ◽

2021 ◽

Vol 14 (4) ◽

pp. 101033

Author(s):

Chia Yin Goh ◽

Sean Patmore ◽

Albert Smolenski ◽

Jane Howard ◽

Shane Evans ◽

...

Keyword(s):

Breast Cancer ◽

Von Willebrand Factor ◽

Cancer Metastasis ◽

Breast Cancer Metastasis ◽

Von Willebrand ◽

Willebrand Factor

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Role of Platelet Membrane Glycoproteins and Von Willebrand Factor in Adhesion of Platelets to Subendothelium and Collagen

Annals of the New York Academy of Sciences ◽

10.1111/j.1749-6632.1987.tb33029.x ◽

1987 ◽

Vol 516 (1 Blood in Cont) ◽

pp. 52-65 ◽

Cited By ~ 33

Author(s):

KJELL S. SAKARIASSEN ◽

EDITH FRESSINAUD ◽

JEAN-PIERRE GIRMA ◽

DOMINIQUE MEYER ◽

HANS R. BAUMGARTNER

Keyword(s):

Von Willebrand Factor ◽

Platelet Membrane ◽

Membrane Glycoproteins ◽

Von Willebrand ◽

Willebrand Factor ◽

Adhesion Of Platelets

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The ADAMTS13 metalloprotease domain: roles of subsites in enzyme activity and specificity

Blood ◽

10.1182/blood-2009-12-258780 ◽

2010 ◽

Vol 116 (16) ◽

pp. 3064-3072 ◽

Cited By ~ 30

Author(s):

Rens de Groot ◽

David A. Lane ◽

James T. B. Crawley

Keyword(s):

Single Point ◽

Minor Importance ◽

Variable Regions ◽

Cleavage Efficiency ◽

Scissile Bond ◽

A Minor ◽

Von Willebrand ◽

A2 Domain ◽

Willebrand Factor

Abstract ADAMTS13 modulates von Willebrand factor (VWF) platelet-tethering function by proteolysis of the Tyr1605-Met1606 bond in the VWF A2 domain. To examine the role of the metalloprotease domain of ADAMTS13 in scissile bond specificity, we identified 3 variable regions (VR1, -2, and -3) in the ADAMTS family metalloprotease domain that flank the active site, which might be important for specificity. Eight composite sequence swaps (to residues in ADAMTS1 or ADAMTS2) and 18 single-point mutants were generated in these VRs and expressed. Swapping VR1 (E184-R193) of ADAMTS13 with that of ADAMTS1 or ADAMTS2 abolished/severely impaired ADAMTS13 function. Kinetic analysis of VR1 point mutants using VWF115 as a short substrate revealed reduced proteolytic function (kcat/Km reduced by 2- to 10-fold) as a result of D187A, R190A, and R193A substitutions. Analysis of VR2 (F216-V220) revealed a minor importance of this region. Mutants of VR3 (G236-A261) proteolysed wild-type VWF115 normally. However, using either short or full-length VWF substrates containing the P1′ M1606A mutation, we identified residues within VR3 (D252-P256) that influence P1′ amino acid specificity, we hypothesize, by shaping the S1′ pocket. It is concluded that 2 subsites, D187-R193 and D252-P256, in the metalloprotease domain play an important role in cleavage efficiency and site specificity.

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