Conformational plasticity of ADAMTS13 in hemostasis and autoimmunity
2021; Elsevier BV; Volume: 297; Issue: 4 Linguagem: Inglês
10.1016/j.jbc.2021.101132
ISSN1083-351X
AutoresBogac Ercig, Tom Arfman, Johana Hrdinová, Kanin Wichapong, Chris Reutelingsperger, Karen Vanhoorelbeke, Gerry A. F. Nicolaes, Jan Voorberg,
Tópico(s)Platelet Disorders and Treatments
ResumoA disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13 (ADAMTS13) is a multidomain metalloprotease for which until now only a single substrate has been identified. ADAMTS13 cleaves the polymeric force-sensor von Willebrand factor (VWF) that unfolds under shear stress and recruits platelets to sites of vascular injury. Shear force–dependent cleavage at a single Tyr–Met peptide bond in the unfolded VWF A2 domain serves to reduce the size of VWF polymers in circulation. In patients with immune-mediated thrombotic thrombocytopenic purpura (iTTP), a rare life-threatening disease, ADAMTS13 is targeted by autoantibodies that inhibit its activity or promote its clearance. In the absence of ADAMTS13, VWF polymers are not adequately processed, resulting in spontaneous adhesion of blood platelets, which presents as severe, life-threatening microvascular thrombosis. In healthy individuals, ADAMTS13–VWF interactions are guided by controlled conversion of ADAMTS13 from a closed, inactive to an open, active conformation through a series of interdomain contacts that are now beginning to be defined. Recently, it has been shown that ADAMTS13 adopts an open conformation in the acute phase and during subclinical disease in iTTP patients, making open ADAMTS13 a novel biomarker for iTTP. In this review, we summarize our current knowledge on ADAMTS13 conformation and speculate on potential triggers inducing conformational changes of ADAMTS13 and how these relate to the pathogenesis of iTTP. A disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13 (ADAMTS13) is a multidomain metalloprotease for which until now only a single substrate has been identified. ADAMTS13 cleaves the polymeric force-sensor von Willebrand factor (VWF) that unfolds under shear stress and recruits platelets to sites of vascular injury. Shear force–dependent cleavage at a single Tyr–Met peptide bond in the unfolded VWF A2 domain serves to reduce the size of VWF polymers in circulation. In patients with immune-mediated thrombotic thrombocytopenic purpura (iTTP), a rare life-threatening disease, ADAMTS13 is targeted by autoantibodies that inhibit its activity or promote its clearance. In the absence of ADAMTS13, VWF polymers are not adequately processed, resulting in spontaneous adhesion of blood platelets, which presents as severe, life-threatening microvascular thrombosis. In healthy individuals, ADAMTS13–VWF interactions are guided by controlled conversion of ADAMTS13 from a closed, inactive to an open, active conformation through a series of interdomain contacts that are now beginning to be defined. Recently, it has been shown that ADAMTS13 adopts an open conformation in the acute phase and during subclinical disease in iTTP patients, making open ADAMTS13 a novel biomarker for iTTP. In this review, we summarize our current knowledge on ADAMTS13 conformation and speculate on potential triggers inducing conformational changes of ADAMTS13 and how these relate to the pathogenesis of iTTP. The increasing prevalence of autoimmune disorders provides a major challenge for the healthcare system worldwide. Our current understanding of the pathogenesis of several autoimmune disorders is incomplete. Environmental triggers have been linked to the onset of immunity. In addition, specific human leukocyte antigen loci have been identified as a risk factor for a large number of autoimmune disorders (1Dendrou C.A. Petersen J. Rossjohn J. Fugger L. HLA variation and disease.Nat. Rev. Immunol. 2018; 18: 325-339Crossref PubMed Scopus (197) Google Scholar). Post-translational modifications such as citrullination have also been implicated in the pathogenesis of autoimmune disorders (2Curran A.M. Naik P. Giles J.T. Darrah E. PAD enzymes in rheumatoid arthritis: Pathogenic effectors and autoimmune targets.Nat. Rev. Rheumatol. 2020; 16: 301-315Crossref PubMed Scopus (23) Google Scholar). Importantly, conformational changes of proteins resulting in exposure of neoepitopes may trigger the development of pathogenic autoantibodies (3Ludwig R.J. Vanhoorelbeke K. Leypoldt F. Kaya Z. Bieber K. McLachlan S.M. Komorowski L. Luo J. Cabral-Marques O. Hammers C.M. Lindstrom J.M. Lamprecht P. Fischer A. Riemekasten G. Tersteeg C. et al.Mechanisms of autoantibody-induced pathology.Front. Immunol. 2017; 8: 603Crossref PubMed Scopus (197) Google Scholar). Conformational changes in beta-2 glycoprotein 1 have been linked to the antiphospholipid syndrome (4de Laat B. de Groot P.G. Autoantibodies directed against domain I of beta2-glycoprotein I.Curr. Rheumatol. Rep. 2011; 13: 70-76Crossref PubMed Scopus (60) Google Scholar), whereas conformational changes in platelet factor 4 upon its binding to heparin trigger the development of anti-platelet factor 4 antibodies resulting in heparin-induced thrombocytopenia (5Kreimann M. Brandt S. Krauel K. Block S. Helm C.A. Weitschies W. Greinacher A. Delcea M. Binding of anti-platelet factor 4/heparin antibodies depends on the thermodynamics of conformational changes in platelet factor 4.Blood. 2014; 124: 2442-2449Crossref PubMed Scopus (53) Google Scholar). In the current review, we explore how conformational plasticity of the protein a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13 (ADAMTS13) relates to the onset of autoimmune thrombotic thrombocytopenic purpura (TTP). TTP is a rare and life-threatening disease with an annual incidence rate of 1.5 to 6 cases per million adults per year (6Sukumar S. Lämmle B. Cataland S.R. Thrombotic thrombocytopenic purpura: Pathophysiology, diagnosis, and management.J. Clin. Med. 2021; 10: 536Crossref PubMed Scopus (8) Google Scholar), which is characterized by a severe deficiency (<10% activity) of the plasma metalloprotease (MP) ADAMTS13. The acute phase of TTP often presents itself with purpura, fever, neurological manifestations, renal dysfunctions, hemolytic anemia with schistocytes, and thrombocytopenia (7Kremer Hovinga J.A. Coppo P. Lämmle B. Moake J.L. Miyata T. Vanhoorelbeke K. Thrombotic thrombocytopenic purpura.Nat. Rev. Dis. Primers. 2017; 3: 17020Crossref PubMed Scopus (132) Google Scholar). Although TTP can be of congenital origin due to mutations in the ADAMTS13 gene, the vast majority of patients develop immune-mediated TTP (iTTP), an autoimmune disorder in which autoantibodies against ADAMTS13 develop (8Joly B.S. Coppo P. Veyradier A. Thrombotic thrombocytopenic purpura.Blood. 2017; 129: 2836-2846Crossref PubMed Scopus (231) Google Scholar). Symptoms of TTP arise because of a patient's inability to cleave ultralarge von Willebrand factor (ULVWF) multimers on the surface of endothelial cells (9Dong J.F. Moake J.L. Nolasco L. Bernardo A. Arceneaux W. Shrimpton C.N. Schade A.J. McIntire L.V. Fujikawa K. López J.A. ADAMTS-13 rapidly cleaves newly secreted ultralarge von Willebrand factor multimers on the endothelial surface under flowing conditions.Blood. 2002; 100: 4033-4039Crossref PubMed Scopus (677) Google Scholar, 10Motto D.G. Chauhan A.K. Zhu G. Homeister J. Lamb C.B. Desch K.C. Zhang W. Tsai H.M. Wagner D.D. Ginsburg D. Shigatoxin triggers thrombotic thrombocytopenic purpura in genetically susceptible ADAMTS13-deficient mice.J. Clin. Invest. 2005; 115: 2752-2761Crossref PubMed Scopus (258) Google Scholar) (Fig. 1). Multimeric von Willebrand factor (VWF) is produced by vascular endothelium and megakaryocytes (11Wagner D.D. Marder V.J. Biosynthesis of von Willebrand protein by human endothelial cells: Processing steps and their intracellular localization.J. Cell Biol. 1984; 99: 2123-2130Crossref PubMed Scopus (204) Google Scholar, 12Sadler J.E. von Willebrand factor.J. Biol. Chem. 1991; 266: 22777-22780Abstract Full Text PDF PubMed Google Scholar). VWF is stored in endothelial cell granules called Weibel–Palade bodies (13Wagner D.D. Olmsted J.B. Marder V.J. Immunolocalization of von Willebrand protein in Weibel-Palade bodies of human endothelial cells.J. Cell Biol. 1982; 95: 355-360Crossref PubMed Scopus (524) Google Scholar), which fuse with the cell membrane to release ULVWF multimers (14Karampini E. Bürgisser P.E. Olins J. Mulder A.A. Jost C.R. Geerts D. Voorberg J. Bierings R. Sec22b determines Weibel-Palade body length by controlling anterograde ER-Golgi transport.Haematologica. 2021; 106: 1138-1147Crossref PubMed Scopus (5) Google Scholar). The ability of VWF to bind circulating platelets is critically dependent on its multimeric size, with the largest multimers being most potent in capturing platelets (15Leebeek F.W.G. Eikenboom J.C.J. Von Willebrand's disease.N. Engl. J. Med. 2017; 376: 701-702Crossref PubMed Scopus (20) Google Scholar) during primary hemostasis. In the absence of ADAMTS13, ULVWF multimers are not processed and platelet-rich clots spontaneously form in the microcirculation, resulting in thrombotic microangiopathy. VWF multimer digestion by ADAMTS13 is dependent on shear stress, which causes the VWF protein overall shape to change from mostly globular to an elongated, stretched form (16Zhang X. Halvorsen K. Zhang C.Z. Wong W.P. Springer T.A. Mechanoenzymatic cleavage of the ultralarge vascular protein von Willebrand factor.Science. 2009; 324: 1330-1334Crossref PubMed Scopus (375) Google Scholar). Upon binding to its substrate VWF, ADAMTS13 adopts a short-lived, transient "open" conformation that allows for multiple exosites within the protease, disintegrin, cysteine-rich (Cys-rich), and spacer domains to interact with complementary sites within the unfolded VWF A2 domain, a process that has been named a "molecular zipper" (17Crawley J.T. de Groot R. Xiang Y. Luken B.M. Lane D.A. Unraveling the scissile bond: How ADAMTS13 recognizes and cleaves von Willebrand factor.Blood. 2011; 118: 3212-3221Crossref PubMed Scopus (196) Google Scholar, 18Petri A. Kim H.J. Xu Y. de Groot R. Li C. Vandenbulcke A. Vanhoorelbeke K. Emsley J. Crawley J.T.B. Crystal structure and substrate-induced activation of ADAMTS13.Nat. Commun. 2019; 10: 3781Crossref PubMed Scopus (23) Google Scholar). Importantly, the "open conformation" refers to the ADAMTS13 conformation that evolves upon release of autoinhibitory interactions between the ADAMTS13 C-terminal complement C1r/C1s, Uegf, Bmp1 (CUB) domains and the spacer domain (Fig. 2A). Hence, when the CUB–spacer interactions are intact, ADAMTS13 is in a "closed" conformation (Fig. 2B). The differences in these conformations can be monitored by a specifically developed mAb that can only bind the spacer domain when the protein is in an "open" conformation (19Roose E. Schelpe A.S. Joly B.S. Peetermans M. Verhamme P. Voorberg J. Greinacher A. Deckmyn H. De Meyer S.F. Coppo P. Veyradier A. Vanhoorelbeke K. An open conformation of ADAMTS-13 is a hallmark of acute acquired thrombotic thrombocytopenic purpura.J. Thromb. Haemost. 2018; 16: 378-388Crossref PubMed Scopus (45) Google Scholar, 20Roose E. Schelpe A.S. Tellier E. Sinkovits G. Joly B.S. Dekimpe C. Kaplanski G. Le Besnerais M. Mancini I. Falter T. Von Auer C. Feys H.B. Reti M. Rossmann H. Vandenbulcke A. et al.Open ADAMTS13, induced by antibodies, is a biomarker for subclinical immune-mediated thrombotic thrombocytopenic purpura.Blood. 2020; 136: 353-361PubMed Google Scholar). Previous literature directly correlates "open" conformation ADAMTS13 to increased proteolytic activity (21South K. Luken B.M. Crawley J.T. Phillips R. Thomas M. Collins R.F. Deforche L. Vanhoorelbeke K. Lane D.A. Conformational activation of ADAMTS13.Proc. Natl. Acad. Sci. U. S. A. 2014; 111: 18578-18583Crossref PubMed Scopus (73) Google Scholar). However, subsequent findings indicate that several conformation-controlled binding events between ADAMTS13 and its substrate VWF are required for the generation of a fully catalytically active conformation of the MP domain (18Petri A. Kim H.J. Xu Y. de Groot R. Li C. Vandenbulcke A. Vanhoorelbeke K. Emsley J. Crawley J.T.B. Crystal structure and substrate-induced activation of ADAMTS13.Nat. Commun. 2019; 10: 3781Crossref PubMed Scopus (23) Google Scholar). This sophisticated sequence of conformational events serves to regulate activation of ADAMTS13 as well as prevent excessive or off-target proteolysis (22South K. Freitas M.O. Lane D.A. Conformational quiescence of ADAMTS-13 prevents proteolytic promiscuity.J. Thromb. Haemost. 2016; 14: 2011-2022Crossref PubMed Scopus (17) Google Scholar). Recently, it was shown that ADAMTS13 circulates in an "open" conformation during the acute phase and during subclinical disease in remission of iTTP, suggesting that a sustained "open" conformation of ADAMTS13 is linked to the onset and recurrence of autoimmunity in these patients (19Roose E. Schelpe A.S. Joly B.S. Peetermans M. Verhamme P. Voorberg J. Greinacher A. Deckmyn H. De Meyer S.F. Coppo P. Veyradier A. Vanhoorelbeke K. An open conformation of ADAMTS-13 is a hallmark of acute acquired thrombotic thrombocytopenic purpura.J. Thromb. Haemost. 2018; 16: 378-388Crossref PubMed Scopus (45) Google Scholar, 20Roose E. Schelpe A.S. Tellier E. Sinkovits G. Joly B.S. Dekimpe C. Kaplanski G. Le Besnerais M. Mancini I. Falter T. Von Auer C. Feys H.B. Reti M. Rossmann H. Vandenbulcke A. et al.Open ADAMTS13, induced by antibodies, is a biomarker for subclinical immune-mediated thrombotic thrombocytopenic purpura.Blood. 2020; 136: 353-361PubMed Google Scholar). Furthermore, in iTTP, conformational changes of ADAMTS13 are observed earlier than detectable titers of anti-ADAMTS13 autoantibodies during an acute episode of iTTP (20Roose E. Schelpe A.S. Tellier E. Sinkovits G. Joly B.S. Dekimpe C. Kaplanski G. Le Besnerais M. Mancini I. Falter T. Von Auer C. Feys H.B. Reti M. Rossmann H. Vandenbulcke A. et al.Open ADAMTS13, induced by antibodies, is a biomarker for subclinical immune-mediated thrombotic thrombocytopenic purpura.Blood. 2020; 136: 353-361PubMed Google Scholar). This observation raises the question how conversion of ADAMTS13 from a closed to an open conformation is regulated and how it relates to the pathogenesis of iTTP. The pathological triggers inducing a switch from the closed to open conformation of ADAMTS13 are now beginning to be defined. For example, it was recently demonstrated that patient-derived anti-ADAMTS13 autoantibodies can promote conversion of ADAMTS13 from a closed to an open conformation (20Roose E. Schelpe A.S. Tellier E. Sinkovits G. Joly B.S. Dekimpe C. Kaplanski G. Le Besnerais M. Mancini I. Falter T. Von Auer C. Feys H.B. Reti M. Rossmann H. Vandenbulcke A. et al.Open ADAMTS13, induced by antibodies, is a biomarker for subclinical immune-mediated thrombotic thrombocytopenic purpura.Blood. 2020; 136: 353-361PubMed Google Scholar, 23Roose E. Veyradier A. Vanhoorelbeke K. Insights into ADAMTS13 structure: Impact on thrombotic thrombocytopenic purpura diagnosis and management.Curr. Opin. Hematol. 2020; 27: 320-326Crossref PubMed Scopus (4) Google Scholar). Apart from autoantibodies, other pathological triggers that have not yet been identified may induce conformational changes of ADAMTS13 in patients with iTTP. In this review, we summarize the impact of key conformational changes that contribute to development of autoimmunity in patients with iTTP. ADAMTS family members are multidomain proteins. They consist of a signal sequence, a propeptide, a metalloproteinase, a disintegrin-like (Dis), a central thrombospondin type 1-like (TSP), a Cys-rich, and a spacer domain and a set of additional TSP repeats (24Kelwick R. Desanlis I. Wheeler G.N. Edwards D.R. The ADAMTS (a disintegrin and metalloproteinase with thrombospondin motifs) family.Genome Biol. 2015; 16: 113Crossref PubMed Scopus (322) Google Scholar) (Fig. 2C). ADAMTS13 uniquely contains two CUB domains at its carboxy terminus (24Kelwick R. Desanlis I. Wheeler G.N. Edwards D.R. The ADAMTS (a disintegrin and metalloproteinase with thrombospondin motifs) family.Genome Biol. 2015; 16: 113Crossref PubMed Scopus (322) Google Scholar). Within the propeptide, a furin-cleavage site is localized which, when cleaved, facilitates removal of the propeptide. ADAMTS MP domains contain a zinc-binding motif with the reprolysin signature (HEXXH + HD). Furthermore, ADAMTS proteins have a conserved cysteine network inside their Cys-rich domain (24Kelwick R. Desanlis I. Wheeler G.N. Edwards D.R. The ADAMTS (a disintegrin and metalloproteinase with thrombospondin motifs) family.Genome Biol. 2015; 16: 113Crossref PubMed Scopus (322) Google Scholar), which is involved in substrate binding in the case of ADAMTS13 (25de Groot R. Lane D.A. Crawley J.T. The role of the ADAMTS13 cysteine-rich domain in VWF binding and proteolysis.Blood. 2015; 125: 1968-1975Crossref PubMed Scopus (27) Google Scholar). The primary role of the ADAMTS13 central TSP repeat seems to be spatial positioning of the VWF-binding exosites present in the surrounding domains (26Xiang Y. de Groot R. Crawley J.T.B. Lane D.A. Mechanism of von Willebrand factor scissile bond cleavage by a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13 (ADAMTS13).Proc. Natl. Acad. Sci. U. S. A. 2011; 108: 11602-11607Crossref PubMed Scopus (38) Google Scholar). Since the isolation of ADAMTS13 from plasma (27Furlan M. Robles R. Lämmle B. Partial purification and characterization of a protease from human plasma cleaving von Willebrand factor to fragments produced by in vivo proteolysis.Blood. 1996; 87: 4223-4234Crossref PubMed Google Scholar, 28Fujikawa K. Suzuki H. McMullen B. Chung D. Purification of human von Willebrand factor-cleaving protease and its identification as a new member of the metalloproteinase family.Blood. 2001; 98: 1662-1666Crossref PubMed Scopus (491) Google Scholar, 29Tsai H.M. Physiologic cleavage of von Willebrand factor by a plasma protease is dependent on its conformation and requires calcium ion.Blood. 1996; 87: 4235-4244Crossref PubMed Google Scholar) and publication of its amino acid sequence in 2001 (28Fujikawa K. Suzuki H. McMullen B. Chung D. Purification of human von Willebrand factor-cleaving protease and its identification as a new member of the metalloproteinase family.Blood. 2001; 98: 1662-1666Crossref PubMed Scopus (491) Google Scholar, 30Gerritsen H.E. Robles R. Lämmle B. Furlan M. Partial amino acid sequence of purified von Willebrand factor-cleaving protease.Blood. 2001; 98: 1654-1661Crossref PubMed Scopus (304) Google Scholar), researchers have intensively focused on understanding the mechanism by which ADAMTS13 processes VWF multimers. The experimental determination of the three-dimensional structure of the disintegrin-like domain, thrombospondin type 1 repeat, cysteine-rich and spacer domains of ADAMTS13 provided a framework for these studies (31Akiyama M. Takeda S. Kokame K. Takagi J. Miyata T. Crystal structures of the noncatalytic domains of ADAMTS13 reveal multiple discontinuous exosites for von Willebrand factor.Proc. Natl. Acad. Sci. U. S. A. 2009; 106: 19274-19279Crossref PubMed Scopus (124) Google Scholar). The crystal structure of the MP domain of ADAMTS13 as well as the crystal structure of the CUB domains were published more recently (18Petri A. Kim H.J. Xu Y. de Groot R. Li C. Vandenbulcke A. Vanhoorelbeke K. Emsley J. Crawley J.T.B. Crystal structure and substrate-induced activation of ADAMTS13.Nat. Commun. 2019; 10: 3781Crossref PubMed Scopus (23) Google Scholar, 32Kim H.J. Xu Y. Petri A. Vanhoorelbeke K. Crawley J.T.B. Emsley J. Crystal structure of ADAMTS13 CUB domains reveals their role in global latency.Sci. Adv. 2021; 7eabg4403Crossref PubMed Scopus (3) Google Scholar). C-terminal domains of ADAMTS family members have been distinguished as important regulators of enzymatic activity (33Tortorella M. Pratta M. Liu R.Q. Abbaszade I. Ross H. Burn T. Arner E. The thrombospondin motif of aggrecanase-1 (ADAMTS-4) is critical for aggrecan substrate recognition and cleavage.J. Biol. Chem. 2000; 275: 25791-25797Abstract Full Text Full Text PDF PubMed Scopus (168) Google Scholar, 34Kashiwagi M. Enghild J.J. Gendron C. Hughes C. Caterson B. Itoh Y. Nagase H. Altered proteolytic activities of ADAMTS-4 expressed by C-terminal processing.J. Biol. Chem. 2004; 279: 10109-10119Abstract Full Text Full Text PDF PubMed Scopus (182) Google Scholar, 35Gendron C. Kashiwagi M. Lim N.H. Enghild J.J. Thøgersen I.B. Hughes C. Caterson B. Nagase H. Proteolytic activities of human ADAMTS-5: Comparative studies with ADAMTS-4.J. Biol. Chem. 2007; 282: 18294-18306Abstract Full Text Full Text PDF PubMed Scopus (199) Google Scholar, 36Flannery C.R. Zeng W. Corcoran C. Collins-Racie L.A. Chockalingam P.S. Hebert T. Mackie S.A. McDonagh T. Crawford T.K. Tomkinson K.N. LaVallie E.R. Morris E.A. Autocatalytic cleavage of ADAMTS-4 (Aggrecanase-1) reveals multiple glycosaminoglycan-binding sites.J. Biol. Chem. 2002; 277: 42775-42780Abstract Full Text Full Text PDF PubMed Scopus (126) Google Scholar, 37Fushimi K. Troeberg L. Nakamura H. Lim N.H. Nagase H. Functional differences of the catalytic and non-catalytic domains in human ADAMTS-4 and ADAMTS-5 in aggrecanolytic activity.J. Biol. Chem. 2008; 283: 6706-6716Abstract Full Text Full Text PDF PubMed Scopus (67) Google Scholar). Importantly, the C-terminal CUB domains are a unique feature of ADAMTS13 and have evolved as critical regulators of ADAMTS13 conformation and activity. Incubation of ADAMTS13 with the anti-CUB2 mAb 20E9 resulted in an increased activity of ADAMTS13 (21South K. Luken B.M. Crawley J.T. Phillips R. Thomas M. Collins R.F. Deforche L. Vanhoorelbeke K. Lane D.A. Conformational activation of ADAMTS13.Proc. Natl. Acad. Sci. U. S. A. 2014; 111: 18578-18583Crossref PubMed Scopus (73) Google Scholar). A similar effect was observed after the addition of the D4-CK fragment derived from VWF (residues 1874–2813), which comprises the natural binding site of the VWF substrate for the ADAMTS13 CUB domains (21South K. Luken B.M. Crawley J.T. Phillips R. Thomas M. Collins R.F. Deforche L. Vanhoorelbeke K. Lane D.A. Conformational activation of ADAMTS13.Proc. Natl. Acad. Sci. U. S. A. 2014; 111: 18578-18583Crossref PubMed Scopus (73) Google Scholar). In addition, mAbs directed toward the T2-T8 repeats and the CUB1/2 domains were also capable of increasing the activity of ADAMTS13 for the fluorescence resonance energy transfer substrate VWF73 (FRETS-VWF73) derived from the A2 domain of the VWF (38Deforche L. Roose E. Vandenbulcke A. Vandeputte N. Feys H.B. Springer T.A. Mi L.Z. Muia J. Sadler J.E. Soejima K. Rottensteiner H. Deckmyn H. De Meyer S.F. Vanhoorelbeke K. Linker regions and flexibility around the metalloprotease domain account for conformational activation of ADAMTS-13.J. Thromb. Haemost. 2015; 13: 2063-2075Crossref PubMed Scopus (39) Google Scholar, 39Schelpe A.-S. Petri A. Roose E. Pareyn I. Deckmyn H. De Meyer S.F. Crawley J.T.B. Vanhoorelbeke K. Antibodies that conformationally activate ADAMTS13 allosterically enhance metalloprotease domain function.Blood Adv. 2020; 4: 1072-1080Crossref PubMed Scopus (14) Google Scholar). These findings suggested that binding of mAbs to the carboxy-terminal TSP2-8 and CUB1-2 domains induces a conformational change in ADAMTS13 that allows for more efficient processing of unfolded peptide substrates such as FRETS-VWF73. This hypothesis is strengthened further by the finding that targeting of the spacer domain by antibodies yields similar results (39Schelpe A.-S. Petri A. Roose E. Pareyn I. Deckmyn H. De Meyer S.F. Crawley J.T.B. Vanhoorelbeke K. Antibodies that conformationally activate ADAMTS13 allosterically enhance metalloprotease domain function.Blood Adv. 2020; 4: 1072-1080Crossref PubMed Scopus (14) Google Scholar). Transmission electron microscopy of negatively stained ADAMTS13 provided evidence for being primarily present in a closed conformation, whereas a so-called gain-of-function variant in which residues Arg568, Phe592, Arg660, Tyr661, and Tyr665 were replaced for corresponding conservative residues was primarily present in an open conformation (21South K. Luken B.M. Crawley J.T. Phillips R. Thomas M. Collins R.F. Deforche L. Vanhoorelbeke K. Lane D.A. Conformational activation of ADAMTS13.Proc. Natl. Acad. Sci. U. S. A. 2014; 111: 18578-18583Crossref PubMed Scopus (73) Google Scholar, 40Jian C. Xiao J. Gong L. Skipwith C.G. Jin S.Y. Kwaan H.C. Zheng X.L. Gain-of-function ADAMTS13 variants that are resistant to autoantibodies against ADAMTS13 in patients with acquired thrombotic thrombocytopenic purpura.Blood. 2012; 119: 3836-3843Crossref PubMed Scopus (86) Google Scholar). Small-angle X-ray scattering combined with molecular modeling provided additional evidence for intradomain interactions between the TSP8-CUB1/2 and the spacer domain (41Muia J. Zhu J. Gupta G. Haberichter S.L. Friedman K.D. Feys H.B. Deforche L. Vanhoorelbeke K. Westfield L.A. Roth R. Tolia N.H. Heuser J.E. Sadler J.E. Allosteric activation of ADAMTS13 by von Willebrand factor.Proc. Natl. Acad. Sci. U. S. A. 2014; 111: 18584-18589Crossref PubMed Scopus (77) Google Scholar). Follow-up studies revealed that both the CUB1 and CUB2 domains are capable of interacting with the spacer domain (Fig. 2B) (42South K. Freitas M.O. Lane D.A. A model for the conformational activation of the structurally quiescent metalloprotease ADAMTS13 by von Willebrand factor.J. Biol. Chem. 2017; 292: 5760-5769Abstract Full Text Full Text PDF PubMed Scopus (23) Google Scholar). A crucial role for TSP8 was suggested in maintaining ADAMTS13 in a closed conformation, most likely by promoting CUB1 spacer domain interactions (42South K. Freitas M.O. Lane D.A. A model for the conformational activation of the structurally quiescent metalloprotease ADAMTS13 by von Willebrand factor.J. Biol. Chem. 2017; 292: 5760-5769Abstract Full Text Full Text PDF PubMed Scopus (23) Google Scholar). In a recent study, Muia et al. showed that most carboxy/terminal domains with the exception of TSP3/TSP6 are needed for allosteric regulation of ADAMTS13 activity (43Zhu J. Muia J. Gupta G. Westfield L.A. Vanhoorelbeke K. Tolia N.H. Sadler J.E. Exploring the "minimal" structure of a functional ADAMTS13 by mutagenesis and small-angle X-ray scattering.Blood. 2019; 133: 1909-1918Crossref PubMed Google Scholar, 44Muia J. Zhu J. Greco S.C. Vanhoorelbeke K. Gupta G. Westfield L.A. Sadler J.E. Phylogenetic and functional analysis of ADAMTS13 identifies highly conserved domains essential for allosteric regulation.Blood. 2019; 133: 1899-1908Crossref PubMed Scopus (15) Google Scholar). Deletion of TSP7 and TSP8 abrogated allosteric regulation of ADAMTS13 (44Muia J. Zhu J. Greco S.C. Vanhoorelbeke K. Gupta G. Westfield L.A. Sadler J.E. Phylogenetic and functional analysis of ADAMTS13 identifies highly conserved domains essential for allosteric regulation.Blood. 2019; 133: 1899-1908Crossref PubMed Scopus (15) Google Scholar). Small-angle X-ray scattering combined with molecular modeling suggested that ADAMTS13 is present in a hairpin-like conformation with a so-called "hinge-like" region between TSP4 and TSP5 (Fig. 2B) (43Zhu J. Muia J. Gupta G. Westfield L.A. Vanhoorelbeke K. Tolia N.H. Sadler J.E. Exploring the "minimal" structure of a functional ADAMTS13 by mutagenesis and small-angle X-ray scattering.Blood. 2019; 133: 1909-1918Crossref PubMed Google Scholar). This finding is consistent with the presence of a linker region between TSP4 and TSP5 that may allow for bending of this part of ADAMTS13 while securing the more rigid disulfide-bonded configuration of the neighboring TSP4 and TSP5 repeats. In agreement with this, an important role for three linker regions after the TSP2, TSP4, and TSP8 domains was proposed to control ADAMTS13 conformation (38Deforche L. Roose E. Vandenbulcke A. Vandeputte N. Feys H.B. Springer T.A. Mi L.Z. Muia J. Sadler J.E. Soejima K. Rottensteiner H. Deckmyn H. De Meyer S.F. Vanhoorelbeke K. Linker regions and flexibility around the metalloprotease domain account for conformational activation of ADAMTS-13.J. Thromb. Haemost. 2015; 13: 2063-2075Crossref PubMed Scopus (39) Google Scholar). Somewhat unexpected, deletion of TSP3/6 repeats in human ADAMTS13 appeared to preserve its ability to convert from a closed to an open conformation (44Muia J. Zhu J. Greco S.C. Vanhoorelbeke K. Gupta G. Westfield L.A. Sadler J.E. Phylogenetic and functional analysis of ADAMTS13 identifies highly conserved domains essential for allosteric regulation.Blood. 2019; 133: 1899-1908Crossref PubMed Scopus (15) Google Scholar). This configuration was derived from a phylogenetic screen, which showed that pigeon ADAMTS13, which lacks the TSP3/TSP6 repeats, is still subject to allosteric regulation (44Muia J. Zhu J. Greco S.C. Vanhoorelbeke K. Gupta G. Westfield L.A. Sadler J.E. Phylogenetic and functional analysis of ADAMTS13 identifies highly conserved domains essential for allosteric regulation.Blood. 2019; 133: 1899-1908Crossref PubMed Scopus (15) Google Scholar). To summarize, ADAMTS13 normally circulates in an autoinhibitory conformation governed by an interaction between its C-terminal domains and the spacer domain which is designated as "closed" conformation. To allow for enzymatic activity, conformational changes in ADAMTS13 are required, which will be addressed below. As previously introduced, multimeric VWF circulates in a globular form; however, exposure to shear forces and the binding to exposed collagen at the site of a damaged vessel promotes its unfolding (45Fu H. Jiang Y. Yang D. Scheiflinger F. Wong W.P. Springer T.A. Flow-induced elongation of von Willebrand factor precedes tension-dependent activation.Nat. Commun. 2017; 8: 324Crossref PubMed Scopus (84) Google Scholar). Single-molecule experiments using optical tweezers have provided evidence for shear force–induced unfolding of VWF that is needed for efficient cleavage of VWF polymers into smaller entities (46Zhang Q. Zhou Y.F. Zhang C.Z. Zhang X. Lu C. Springer T.A. Structural specializations of A2, a force-sensing
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