Could shear stress be the answer?
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《血液学杂志》
This study demonstrates that mutations within the A2 domain that impair the secretion of VWF also influence the proteolysis of VWF by ADAMTS13 because this effect is a general property of VWD type 2A.
Von Willebrand factor (VWF) is a large multimeric glycoprotein that mediates platelet adhesion under conditions of elevated fluid shear stress. The mature polypeptide of VWF consists of 2050 amino acids, and it first dimerizes then polymerizes to generate multimers that may exceed 20 000 kDa. VWF is synthesized by megakaryocytes and endothelial cells. The latter secrete ultralarge multimers of VWF into plasma, where they are cleaved by the metalloprotease ADAMTS13, which cleaves at a single site within the second of 3 consecutive A domains (the A2 domain), between residues Y1605 and M1606. The high-molecular-weight multimers (HMWMs) of VWF are the most active in hemostasis. This fact is supported by the naturally occurring mutations that cause von Willebrand disease (VWD) type 2A. The bleeding observed in this condition is due to the loss of the HMWMs of VWF, and most mutations have been identified within the A2 domain of VWF. These mutations have been classified as group 1, the mutations that impair the secretion of VWF into plasma, while group 2 represents those that enhance proteolysis by ADAMTS13.
In this issue of Blood, Hassenpflug and colleagues demonstrate that type 2A mutations of both groups 1 and 2 enhance the susceptibility to proteolysis by ADAMTS13. To evaluate the effect of the mutations, the authors examined the ability of recombinant ADAMTS13 to cleave recombinant full-length VWF variants containing type 2A mutations of both groups. This approach precluded the use of plasma as a source of ADAMTS13, and the results were influenced only by the enzyme and its substrate. Eleven of the 13 VWF mutants tested displayed enhanced susceptibility to proteolysis, and the authors demonstrate for the first time that 3 mutations known to impair secretion of VWF (group 1) also increased its susceptibility to proteolysis by ADAMTS13. Thus, the authors suggest that any mutation within the A2 domain of VWF that increases the susceptibility to proteolysis by ADAMTS13 is a general property of VWD type 2A.
It is thus possible that mutations within the A2 domain may act not only by destabilizing the overall structure of the domain, but could weaken its interaction with either or both of the other A domains, A1 or A3. In both cases the effect seems to be independent of perturbing the tri-domain conformation under stasis, because the authors observed that the chaotropic agent urea was not sufficient to increase proteolysis in some mutants. As acknowledged by the authors, shear forces play an important role on the activity of ADAMTS13 in vivo,1 and thus, it is most likely that the type 2A mutants make the A domain(s) more susceptible to shear-induced conformational change. This could explain why in this study, which was carried out under static conditions, 2 mutants (G1505E and I1628T) failed to enhance susceptibility to proteolysis. The notion that shear stress is required for efficient VWF cleavage is supported by the observations that a recombinant A2 domain protein is rapidly cleaved in the absence of both denaturant agents and divalent cations2 and that recombinant A2 variants containing type 2A mutations of group 2 had little or no effect on proteolysis by ADAMTS13.3 As suggested by the authors, high-resolution structures of VWF or of the A1A2A3 region will help us to understand the molecular mechanism by which the A2 domain in VWF is altered to expose the cleavage site for ADAMTS13 in flowing blood.
References
Dong JF, Moake JL, Nolasco L, et al. ADAMTS-13 rapidly cleaves newly secreted ultralarge von Willebrand factor multimers on the endothelial surface under flowing conditions. Blood. 2002;100: 4033-4039.
Cruz MA, Whitelock J, Dong JF. Evaluation of ADAMTS-13 activity in plasma using recombinant von Willebrand Factor A2 domain polypeptide as substrate. Thromb Haemost. 2003;90: 1204-1209.
Zanardelli S, Crawley JT, Chion CK, et al. ADAMTS13 substrate recognition of von Willebrand factor A2 domain. J Biol Chem. Prepublished October 12, 2005, as DOI 10.1074/jbc.M508316200.(Miguel A. Cruz)
Von Willebrand factor (VWF) is a large multimeric glycoprotein that mediates platelet adhesion under conditions of elevated fluid shear stress. The mature polypeptide of VWF consists of 2050 amino acids, and it first dimerizes then polymerizes to generate multimers that may exceed 20 000 kDa. VWF is synthesized by megakaryocytes and endothelial cells. The latter secrete ultralarge multimers of VWF into plasma, where they are cleaved by the metalloprotease ADAMTS13, which cleaves at a single site within the second of 3 consecutive A domains (the A2 domain), between residues Y1605 and M1606. The high-molecular-weight multimers (HMWMs) of VWF are the most active in hemostasis. This fact is supported by the naturally occurring mutations that cause von Willebrand disease (VWD) type 2A. The bleeding observed in this condition is due to the loss of the HMWMs of VWF, and most mutations have been identified within the A2 domain of VWF. These mutations have been classified as group 1, the mutations that impair the secretion of VWF into plasma, while group 2 represents those that enhance proteolysis by ADAMTS13.
In this issue of Blood, Hassenpflug and colleagues demonstrate that type 2A mutations of both groups 1 and 2 enhance the susceptibility to proteolysis by ADAMTS13. To evaluate the effect of the mutations, the authors examined the ability of recombinant ADAMTS13 to cleave recombinant full-length VWF variants containing type 2A mutations of both groups. This approach precluded the use of plasma as a source of ADAMTS13, and the results were influenced only by the enzyme and its substrate. Eleven of the 13 VWF mutants tested displayed enhanced susceptibility to proteolysis, and the authors demonstrate for the first time that 3 mutations known to impair secretion of VWF (group 1) also increased its susceptibility to proteolysis by ADAMTS13. Thus, the authors suggest that any mutation within the A2 domain of VWF that increases the susceptibility to proteolysis by ADAMTS13 is a general property of VWD type 2A.
It is thus possible that mutations within the A2 domain may act not only by destabilizing the overall structure of the domain, but could weaken its interaction with either or both of the other A domains, A1 or A3. In both cases the effect seems to be independent of perturbing the tri-domain conformation under stasis, because the authors observed that the chaotropic agent urea was not sufficient to increase proteolysis in some mutants. As acknowledged by the authors, shear forces play an important role on the activity of ADAMTS13 in vivo,1 and thus, it is most likely that the type 2A mutants make the A domain(s) more susceptible to shear-induced conformational change. This could explain why in this study, which was carried out under static conditions, 2 mutants (G1505E and I1628T) failed to enhance susceptibility to proteolysis. The notion that shear stress is required for efficient VWF cleavage is supported by the observations that a recombinant A2 domain protein is rapidly cleaved in the absence of both denaturant agents and divalent cations2 and that recombinant A2 variants containing type 2A mutations of group 2 had little or no effect on proteolysis by ADAMTS13.3 As suggested by the authors, high-resolution structures of VWF or of the A1A2A3 region will help us to understand the molecular mechanism by which the A2 domain in VWF is altered to expose the cleavage site for ADAMTS13 in flowing blood.
References
Dong JF, Moake JL, Nolasco L, et al. ADAMTS-13 rapidly cleaves newly secreted ultralarge von Willebrand factor multimers on the endothelial surface under flowing conditions. Blood. 2002;100: 4033-4039.
Cruz MA, Whitelock J, Dong JF. Evaluation of ADAMTS-13 activity in plasma using recombinant von Willebrand Factor A2 domain polypeptide as substrate. Thromb Haemost. 2003;90: 1204-1209.
Zanardelli S, Crawley JT, Chion CK, et al. ADAMTS13 substrate recognition of von Willebrand factor A2 domain. J Biol Chem. Prepublished October 12, 2005, as DOI 10.1074/jbc.M508316200.(Miguel A. Cruz)