{"gene":"ADAMTS13","run_date":"2026-04-28T17:12:37","timeline":{"discoveries":[{"year":2001,"finding":"ADAMTS13 (VWF-cleaving protease) was identified as a new member of the ADAMTS family of metalloproteases with a domain architecture including: signal peptide, short propeptide (with furin consensus RQRR), reprolysin-like metalloprotease domain, disintegrin-like domain, thrombospondin-1 repeat, Cys-rich domain, ADAMTS spacer, seven additional thrombospondin-1 repeats, and two CUB domains. The Cys-rich domain contains an RGDS sequence. Sites for Zn2+ and Ca2+ are conserved in the protease domain.","method":"cDNA cloning, Northern blotting, sequence analysis","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 — original cloning paper with full structural characterization, highly cited foundational study","pmids":["11557746"],"is_preprint":false},{"year":2002,"finding":"Recombinant ADAMTS13 expressed in HEK 293 cells degrades VWF multimers and proteolytically cleaves VWF to the same fragments as plasma VWF-cleaving protease, confirming that ADAMTS13 is responsible for physiological proteolytic degradation of VWF multimers. Recombinant ADAMTS13-mediated degradation was entirely inhibited by plasma from a patient with acquired TTP.","method":"Expression of recombinant ADAMTS13 in HEK 293 cells, VWF multimer degradation assay, inhibition by patient plasma","journal":"Blood","confidence":"High","confidence_rationale":"Tier 1 — direct functional reconstitution of enzymatic activity with controls","pmids":["12393399"],"is_preprint":false},{"year":2003,"finding":"The ADAMTS13 propeptide is not required for folding, secretion, or enzymatic activity. Furin is the protease responsible for propeptide cleavage. Secreted pro-ADAMTS13 (with intact propeptide) has normal proteolytic activity toward VWF. In cells co-expressing ADAMTS13 and VWF, pro-ADAMTS13 can cleave pro-VWF intracellularly.","method":"Propeptide deletion and furin consensus site mutagenesis, expression in HeLa and furin-deficient LoVo cells, VWF cleavage assay","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 — mutagenesis plus reconstitution in multiple cell lines with rigorous controls","pmids":["12975358"],"is_preprint":false},{"year":2007,"finding":"ADAMTS13 is O-fucosylated on at least six of its eight thrombospondin type 1 repeats (TSRs) with a glucose-β1,3-fucose disaccharide. This modification is required for efficient secretion of ADAMTS13; mutation of O-fucosylation sites or knockdown of POFUT2 (the transferase responsible) markedly reduces ADAMTS13 secretion.","method":"Mass spectral analysis of tryptic peptides, metabolic labeling with [3H]fucose, site-directed mutagenesis of modified serines, siRNA knockdown of POFUT2, GDP-fucose-deficient cell line","journal":"The Journal of biological chemistry","confidence":"High","confidence_rationale":"Tier 1 — multiple orthogonal methods including MS, mutagenesis, and genetic depletion","pmids":["17395589"],"is_preprint":false},{"year":2010,"finding":"The ADAMTS13 metalloprotease domain contains two subsites critical for VWF cleavage: residues D187-R193 (VR1) are important for cleavage efficiency (kcat/Km reduced 2–10 fold by point mutations D187A, R190A, R193A), and residues D252-P256 (VR3) influence P1' amino acid specificity, shaping the S1' substrate-binding pocket.","method":"Composite sequence swaps and single-point mutagenesis in variable regions; kinetic analysis using VWF115 substrate and P1'-mutated VWF substrates","journal":"Blood","confidence":"High","confidence_rationale":"Tier 1 — systematic mutagenesis with kinetic readout identifies catalytic subsites","pmids":["20647566"],"is_preprint":false},{"year":2011,"finding":"ADAMTS13 proteolysis of VWF is controlled not by inhibitors but by conformational changes in VWF induced by elevated rheological shear forces, which unfold the VWF A2 domain to reveal cryptic exosites and the scissile bond (Tyr1605-Met1606). ADAMTS13 makes multiple exosite interactions with the unfolded A2 domain to position itself for cleavage.","method":"Review synthesizing biochemical and biophysical studies; supported by experimental literature on shear-dependent VWF unfolding and ADAMTS13 exosite mapping","journal":"Blood","confidence":"High","confidence_rationale":"Tier 2 — synthesis of multiple experimental studies with strong mechanistic consensus","pmids":["21715306"],"is_preprint":false},{"year":2008,"finding":"ADAMTS13 catalysis of VWF A2-domain peptide (FRETS-VWF73) cleavage at Tyr1605-Met1606 is governed by two ionizable groups (pKa ~6.4 and ~4.0). The transition state is more ordered than the ground state (negative activation entropy). Residues Asp1653, Glu1655, Glu1660, Asp1663, and Thr1656 in the VWF A2 sequence form a 'hot spot' driving molecular recognition and allosteric regulation of ADAMTS13 binding. The product peptide (Met1606-Arg1668) acts as a hyperbolic mixed-type inhibitor.","method":"Fluorescence quenching (FRETS) kinetic assay across temperature and pH ranges; Co2+-substituted enzyme; product inhibition analysis; molecular modeling","journal":"Biophysical journal","confidence":"High","confidence_rationale":"Tier 1 — detailed in vitro enzymology with multiple physical-chemical approaches","pmids":["18502798"],"is_preprint":false},{"year":2014,"finding":"ADAMTS13 is allosterically regulated: the distal T8-CUB2 domains interact with and inhibit the proximal MDTCS catalytic domains. Binding of VWF D4 domain (or monoclonal antibodies) to the distal domains relieves this autoinhibition and markedly enhances substrate cleavage. Small angle X-ray scattering data support interdomain contact between distal T-CUB and proximal MDTCS domains in the closed, autoinhibited conformation.","method":"VWF D4-CUB binding assay, monoclonal antibody activation, kinetic proteolysis assays with truncation variants, small angle X-ray scattering (SAXS)","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"High","confidence_rationale":"Tier 1 — reconstitution with multiple domain variants, SAXS structural data, replicated with antibody activation","pmids":["25512528"],"is_preprint":false},{"year":2019,"finding":"Crystal structure of ADAMTS13 (metalloprotease to spacer domains) reveals a latent conformation in which the active-site cleft is occluded, requiring allosteric change for substrate accommodation. Kinetic analyses show that cysteine-rich and spacer domain exosites bring enzyme and substrate into proximity, then disintegrin-like domain exosite binding to VWF allosterically activates the adjacent metalloprotease domain. VWF functions as both the activating cofactor and substrate.","method":"Crystal structure determination, kinetic analyses of VWF proteolysis by exosite mutants","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 1 — crystal structure combined with kinetic functional validation, multiple orthogonal approaches","pmids":["31439947"],"is_preprint":false},{"year":2019,"finding":"ADAMTS13 allosteric activation is conserved across vertebrates. The T7 and T8 distal thrombospondin domains are essential for allosteric activation by VWF D4 domain; deletion of T7 or T8 abolishes allosteric activation. Domains T3-T6 are dispensable for allosteric regulation and shear-dependent cleavage.","method":"Assay of ADAMTS13 from 20 placental mammals, birds, and amphibians; phylogenetic analysis; domain deletion mutants tested for VWF D4-induced activation and shear-dependent cleavage","journal":"Blood","confidence":"High","confidence_rationale":"Tier 1 — systematic deletion mutagenesis combined with cross-species analysis, multiple orthogonal assays","pmids":["30700419"],"is_preprint":false},{"year":2020,"finding":"Conformational activation of ADAMTS13 by anti-Spacer (3E4) or anti-CUB1 (17G2) monoclonal antibodies enhances ADAMTS13 catalytic efficiency (kcat/Km) ~2-fold. The mechanism is allosteric enhancement of metalloprotease domain active site function (increased kcat), not exposure of Spacer or cysteine-rich exosites or improved substrate binding (Km unchanged). A cryptic metalloprotease domain epitope is exposed upon mAb-induced conformational extension.","method":"Kinetic analyses of VWF A2 domain fragment proteolysis; ELISA-based conformational assay; kcat and Km determination","journal":"Blood advances","confidence":"High","confidence_rationale":"Tier 1 — kinetic dissection of activation mechanism with two independent mAbs and multiple readouts","pmids":["32196558"],"is_preprint":false},{"year":2005,"finding":"Platelets contain functionally active ADAMTS13. Platelet lysates cleave endothelial cell-derived ultra-large VWF under static and flow conditions in a divalent cation-dependent manner; cleavage is inhibited by EDTA and by anti-ADAMTS13 antibody from a TTP patient. ADAMTS13 surface expression on platelets increases upon activation by thrombin receptor-activating peptide but not ADP.","method":"VWF cleavage assays with platelet lysates, antibody inhibition, flow cytometry, immunoblotting with four domain-specific antibodies","journal":"Journal of thrombosis and haemostasis : JTH","confidence":"High","confidence_rationale":"Tier 2 — multiple antibodies, functional assay under flow, specific inhibition controls","pmids":["16176307"],"is_preprint":false},{"year":2011,"finding":"Glomerular endothelial cells express and secrete functionally active ADAMTS13, which cleaves VWF. ADAMTS13 deficiency in mice results in thickening of glomerular capillaries with platelet deposition on the vessel wall.","method":"Real-time PCR, flow cytometry, immunofluorescence, immunoblotting, VWF multimer analysis in human glomerular endothelial cells; immunohistochemistry and electron microscopy in wild-type vs. Adamts13−/− mice","journal":"PloS one","confidence":"High","confidence_rationale":"Tier 2 — multiple orthogonal methods in both human cells and genetic mouse model with functional readout","pmids":["21720563"],"is_preprint":false},{"year":2009,"finding":"ADAMTS13 deficiency aggravates ischemic brain damage in mice via the VWF-platelet axis: after reperfusion, Adamts13−/− mice show markedly decreased regional cerebral blood flow and larger infarct volumes compared to wild-type mice, demonstrating that ADAMTS13 protects against ischemia-reperfusion injury by regulating VWF-platelet interactions.","method":"Middle cerebral artery occlusion model in Adamts13−/− vs. wild-type mice; cerebral blood flow measurement; infarct volume quantification","journal":"Blood","confidence":"High","confidence_rationale":"Tier 2 — genetic knockout model with defined mechanistic phenotype (VWF-platelet axis) and quantitative endpoints","pmids":["19965676"],"is_preprint":false},{"year":2013,"finding":"The N-terminal MDTCS portion of ADAMTS13 is necessary and sufficient for proteolytic cleavage of VWF and attenuation of arterial/venous thrombosis. The distal portion (TSP1 2-8 repeats and CUB domains) may function as a disulfide bond reductase to prevent elongation of ultra-large VWF strings on activated endothelial cells. Proteolytic cleavage of VWF by ADAMTS13 is accelerated by FVIII and platelets under fluid shear stress; disruption of FVIII (or platelet glycoprotein 1bα)–VWF interactions dramatically impairs ADAMTS13-dependent proteolysis in vitro and in vivo.","method":"Domain truncation variants, thrombosis models in Adamts13−/− mice, in vitro VWF cleavage assays under flow","journal":"Journal of thrombosis and haemostasis : JTH","confidence":"High","confidence_rationale":"Tier 2 — structure-function with multiple domain variants and genetic/in vitro validation","pmids":["23809107"],"is_preprint":false},{"year":2014,"finding":"ADAMTS13 modulates atherosclerotic plaque progression through a VWF-dependent mechanism: deficiency of VWF in Adamts13−/−/ApoE−/− triple knockout mice completely reverses exacerbated atherosclerosis, macrophage/neutrophil infiltration, and interstitial collagen content compared with Adamts13−/−/ApoE−/− mice.","method":"Genetic epistasis using Adamts13−/−/Vwf−/−/ApoE−/− triple knockout mice on high-fat diet; aortic sinus cross-section analysis; immunohistochemistry","journal":"Journal of thrombosis and haemostasis : JTH","confidence":"High","confidence_rationale":"Tier 2 — genetic epistasis with triple knockout cleanly placing ADAMTS13 upstream of VWF in atherosclerosis","pmids":["24261607"],"is_preprint":false},{"year":2012,"finding":"Recombinant full-length ADAMTS13 promotes endothelial tube formation, proliferation, and migration in a dose-dependent manner independent of VEGF. ADAMTS13 also inhibits VEGF-induced angiogenesis, and these effects are mediated through the C-terminal TSP1 domains. Co-immunoprecipitation shows ADAMTS13 binds to VEGF via its TSP1 domain.","method":"Matrigel tube formation assay, proliferation counting assay, Boyden chamber migration assay, antibody blockade, truncation variants, co-immunoprecipitation","journal":"Microvascular research","confidence":"Medium","confidence_rationale":"Tier 2 — multiple functional assays with domain mapping and co-IP, but single lab","pmids":["22626948"],"is_preprint":false},{"year":2014,"finding":"ADAMTS13 promotes angiogenesis via upregulation of VEGF expression and VEGFR2 phosphorylation in endothelial cells. The C-terminal TSP1 repeats (domains 2-8) are responsible for this angiogenic activity, as shown by structure-function analysis with truncation variants. Anti-VEGF IgG abrogates TSP1-domain-mediated effects on proliferation, migration, and VEGFR2 phosphorylation.","method":"Western blot, ELISA, VEGFR2 phosphorylation assay, proliferation and chemotaxis assays with TSP1-domain truncation variants and anti-VEGF antibody","journal":"Cellular and molecular life sciences : CMLS","confidence":"Medium","confidence_rationale":"Tier 2 — multiple assays with domain mapping and antibody inhibition, single lab","pmids":["24950743"],"is_preprint":false},{"year":2009,"finding":"Autologous transplantation of hematopoietic progenitor cells transduced with lentiviral vector encoding ADAMTS13 restores plasma ADAMTS13 activity in Adamts13−/− mice, reduces VWF multimer size, and significantly prolongs ferric chloride-induced carotid arterial occlusion time, establishing hematopoietic cells as a source of functional ADAMTS13.","method":"Lentiviral transduction of hematopoietic progenitor cells, autologous transplantation in Adamts13−/− mice, plasma ADAMTS13 activity assay, VWF multimer analysis, ferric chloride thrombosis model","journal":"Blood","confidence":"High","confidence_rationale":"Tier 2 — genetic correction with functional endpoint in vivo, multiple readouts","pmids":["19141866"],"is_preprint":false},{"year":2017,"finding":"ADAMTS13 controls vascular remodeling after ischemic stroke by modifying VWF reactivity. ADAMTS13-deficient mice show reduced neovascularization, impaired capillary perfusion, pericyte loss, and accelerated blood-brain barrier breakdown after stroke. VWF deficiency or anti-VWF antibody treatment reversed these deficits in Adamts13−/− mice, placing ADAMTS13 upstream of VWF in post-stroke angiogenesis.","method":"Adamts13−/−, Vwf−/−, and double-knockout mice in stroke model; microvascular analysis; angiopoietin-2 and galectin-3 measurement; adenoviral and recombinant protein rescue experiments; VEGFR2 antagonist","journal":"Blood","confidence":"High","confidence_rationale":"Tier 2 — genetic epistasis with VWF knockout plus multiple rescue experiments establishing pathway","pmids":["28428179"],"is_preprint":false},{"year":2019,"finding":"ADAMTS13 deficiency combined with complement factor H (cfh) heterozygous mutation (cfh R1206) synergistically induces thrombotic microangiopathy in mice, whereas either single defect alone does not cause overt disease, demonstrating epistatic interaction between the ADAMTS13-VWF axis and complement activation in TMA pathogenesis.","method":"Genetic epistasis using Adamts13−/−, cfh-mutant, and double-mutant mice; platelet counts, haptoglobin, erythrocyte fragmentation, LDH, BUN, creatinine, histology, immunohistochemistry","journal":"Blood","confidence":"High","confidence_rationale":"Tier 2 — clean genetic epistasis with quantitative phenotypic endpoints across multiple genotypes","pmids":["31409673"],"is_preprint":false},{"year":2005,"finding":"The transcriptional activity of the ADAMTS13 gene is not significantly altered by proinflammatory stimuli (endotoxin, TNF-α, IL-6, IL-1β), immunosuppressive agents (cyclosporine A, steroids, doxycycline), suggesting that diminished ADAMTS13 activity observed under various clinical conditions is not primarily due to transcriptional regulation.","method":"RNA analysis in liver homogenates and Hep3B liver cell culture; stimulation with multiple agents; in silico promoter analysis","journal":"Thrombosis and haemostasis","confidence":"Medium","confidence_rationale":"Tier 2 — systematic stimulation panel with gene expression readout in relevant liver cell system","pmids":["16113782"],"is_preprint":false},{"year":2015,"finding":"Platelet-delivered recombinant ADAMTS13 (expressed in transgenic platelets) is released upon stimulation with thrombin and collagen, inhibits arterial thrombosis after vascular injury, and prevents TTP syndrome in mice even in the absence of plasma ADAMTS13 activity or in the presence of anti-ADAMTS13 autoantibodies.","method":"Transgenic mice expressing ADAMTS13 in platelets, Western blot, FRET activity assay, ferric chloride thrombosis model, Shigatoxin-2 and VWF-induced TTP models","journal":"Blood","confidence":"High","confidence_rationale":"Tier 2 — proof-of-concept in multiple in vivo models with functional activity assays","pmids":["25800050"],"is_preprint":false},{"year":2012,"finding":"The epitope targeted by inhibitory anti-ADAMTS13 autoantibodies in acquired TTP is primarily within the spacer domain, specifically a surface comprising Arg660, Tyr661, and Tyr665, which contributes to productive binding of ADAMTS13 to unfolded VWF. IgG autoantibodies are predominantly of the IgG4 subclass.","method":"Epitope mapping, patient autoantibody characterization, domain-specific inhibition studies","journal":"Hematology. American Society of Hematology. Education Program","confidence":"Medium","confidence_rationale":"Tier 2 — epitope mapping with functional inhibition data, supported across multiple patient studies","pmids":["23233642"],"is_preprint":false},{"year":2020,"finding":"The immunogenic hotspots in the ADAMTS13 spacer domain targeted by anti-spacer autoantibodies in iTTP are amino acid regions 588-592, 602-610, and 657-666, identified using a library of 11 full-length ADAMTS13 spacer hybrids with corresponding ADAMTS1 spacer regions substituted.","method":"Library of 11 spacer hybrid ADAMTS13 proteins; ELISA epitope mapping with 138 iTTP patient plasma samples","journal":"Journal of thrombosis and haemostasis : JTH","confidence":"High","confidence_rationale":"Tier 2 — systematic epitope mapping with large patient cohort and multiple hybrid constructs","pmids":["33171004"],"is_preprint":false}],"current_model":"ADAMTS13 is a constitutively active plasma metalloprotease that cleaves the Tyr1605-Met1606 bond in the VWF A2 domain under fluid shear stress; its activity is allosterically regulated by autoinhibitory contacts between C-terminal T8-CUB2 domains and the proximal MDTCS catalytic module, which are relieved by VWF D4 domain (or platelet/FVIII cofactor) binding to expose the active site, while the protease is secreted from liver and endothelial cells in a process requiring O-fucosylation of TSR domains, and its deficiency—whether hereditary or due to inhibitory autoantibodies targeting the spacer domain—causes accumulation of ultra-large VWF multimers and thrombotic thrombocytopenic purpura."},"narrative":{"teleology":[{"year":2001,"claim":"Identification of the VWF-cleaving protease as a new ADAMTS family metalloprotease established its domain architecture—metalloprotease, disintegrin-like, TSR, Cys-rich, spacer, seven additional TSRs, and two CUB domains—providing the structural framework for all subsequent functional dissection.","evidence":"cDNA cloning, Northern blotting, and sequence analysis of the human gene","pmids":["11557746"],"confidence":"High","gaps":["No three-dimensional structure yet available","Expression sites and regulation unknown","Catalytic mechanism not characterized"]},{"year":2002,"claim":"Recombinant ADAMTS13 reconstituted VWF multimer degradation identical to the plasma protease and was inhibited by acquired TTP patient plasma, formally proving that ADAMTS13 is the physiological VWF-cleaving protease and that autoantibodies cause its functional deficiency.","evidence":"Recombinant expression in HEK 293 cells with VWF multimer degradation assay and patient-plasma inhibition","pmids":["12393399"],"confidence":"High","gaps":["Cleavage site specificity and kinetics not defined","Propeptide role unknown","In vivo relevance not yet demonstrated in genetic models"]},{"year":2003,"claim":"Demonstrating that the propeptide is dispensable for folding, secretion, and enzymatic activity—and that pro-ADAMTS13 can cleave pro-VWF intracellularly—established that ADAMTS13 is constitutively active, unlike most ADAMTS family members that require propeptide removal for activation.","evidence":"Propeptide deletion and furin-site mutagenesis in HeLa and furin-deficient LoVo cells with VWF cleavage assays","pmids":["12975358"],"confidence":"High","gaps":["Whether intracellular cleavage is physiologically significant in vivo","Regulatory mechanisms controlling activity remain unknown"]},{"year":2005,"claim":"Discovery of functionally active ADAMTS13 in platelets and on the platelet surface (upregulated by thrombin) revealed a local, on-demand source of VWF-cleaving activity at sites of thrombus formation, beyond the known hepatic secretion.","evidence":"Platelet lysate VWF cleavage assay under static and flow conditions, flow cytometry, and inhibition by anti-ADAMTS13 antibody","pmids":["16176307"],"confidence":"High","gaps":["Relative contribution of platelet versus plasma ADAMTS13 in vivo unknown","Mechanism of platelet ADAMTS13 storage and release not defined"]},{"year":2007,"claim":"Identification of O-fucosylation on six of eight TSR domains and its requirement for secretion established a critical post-translational quality-control step: POFUT2-dependent glycosylation is necessary for ADAMTS13 to exit the ER efficiently.","evidence":"Mass spectrometry, metabolic labeling, site-directed mutagenesis of O-fucose sites, and siRNA knockdown of POFUT2","pmids":["17395589"],"confidence":"High","gaps":["Whether O-fucosylation also affects catalytic activity or stability in plasma","Role of the glucose-β1,3-fucose disaccharide versus fucose alone"]},{"year":2008,"claim":"Detailed enzymology of the Tyr1605-Met1606 cleavage reaction—revealing two catalytic ionizable groups, negative activation entropy, and a VWF A2 'hot spot' driving molecular recognition—provided the first kinetic and thermodynamic framework for ADAMTS13 catalysis.","evidence":"FRETS-VWF73 fluorescence quenching kinetics across pH and temperature; Co²⁺-substituted enzyme; product inhibition analysis","pmids":["18502798"],"confidence":"High","gaps":["Structural basis of the two ionizable groups not resolved","Catalytic mechanism at atomic resolution still missing"]},{"year":2009,"claim":"Genetic knockout studies in stroke and gene-therapy models demonstrated that ADAMTS13 deficiency worsens ischemia-reperfusion injury via the VWF-platelet axis and that hematopoietic cell-derived ADAMTS13 can rescue plasma activity, establishing both the in vivo protective role and a potential therapeutic source.","evidence":"Middle cerebral artery occlusion in Adamts13⁻/⁻ mice; lentiviral transduction of hematopoietic progenitor cells with autologous transplantation and thrombosis assays","pmids":["19965676","19141866"],"confidence":"High","gaps":["Contribution of brain endothelial ADAMTS13 versus circulating protease not separated","Optimal therapeutic delivery route undetermined"]},{"year":2010,"claim":"Mutagenesis of the metalloprotease domain identified two substrate-contacting variable regions (VR1: D187-R193 for efficiency; VR3: D252-P256 for P1' specificity), beginning to define the molecular determinants of ADAMTS13's exquisite VWF selectivity.","evidence":"Composite sequence swaps and point mutations with kinetic analysis using VWF115 substrate","pmids":["20647566"],"confidence":"High","gaps":["Structure of the enzyme–substrate complex not yet solved","Full exosite map incomplete"]},{"year":2011,"claim":"Synthesis of biochemical and biophysical evidence established that shear-induced unfolding of the VWF A2 domain—not a classical protease inhibitor—is the primary regulatory mechanism controlling ADAMTS13 proteolysis, and that glomerular endothelial cells are a functionally relevant local source of the enzyme.","evidence":"Review integrating shear-dependent VWF unfolding and exosite mapping studies; real-time PCR, immunofluorescence, and VWF multimer analysis in human glomerular endothelial cells and Adamts13⁻/⁻ mice","pmids":["21715306","21720563"],"confidence":"High","gaps":["Quantitative threshold of shear force needed for A2 domain unfolding in vivo not precisely defined","Relative importance of endothelial versus hepatic ADAMTS13 unclear"]},{"year":2012,"claim":"Mapping the autoantibody epitope in acquired TTP to the spacer domain surface (Arg660/Tyr661/Tyr665) explained how autoantibodies inhibit VWF binding and established the spacer domain as both a critical substrate-recognition exosite and the principal immunogenic target.","evidence":"Epitope mapping with patient autoantibodies, domain-specific inhibition studies, IgG subclass characterization","pmids":["23233642"],"confidence":"Medium","gaps":["Whether epitope escape mutations can be engineered without loss of activity","T-cell epitopes driving the autoimmune response not mapped"]},{"year":2014,"claim":"Discovery of allosteric autoinhibition—where distal T8-CUB2 domains fold back on MDTCS and VWF D4 binding relieves this inhibition—fundamentally reframed ADAMTS13 as a conformationally regulated enzyme rather than a constitutively open protease, and genetic epistasis with VWF deficiency placed it upstream of VWF in atherosclerotic plaque progression.","evidence":"SAXS, domain truncation kinetics, and monoclonal antibody activation; Adamts13⁻/⁻/Vwf⁻/⁻/ApoE⁻/⁻ triple knockout mice on high-fat diet","pmids":["25512528","24261607"],"confidence":"High","gaps":["Full-length structure in open and closed states not yet solved","Whether additional cofactors modulate the open-closed transition in vivo"]},{"year":2019,"claim":"The crystal structure of the MDTCS module revealed a latent active-site conformation requiring allosteric opening, and cross-species analysis showed that T7-T8 domains are specifically essential for VWF D4-mediated allosteric activation, while T3-T6 are dispensable.","evidence":"Crystal structure determination with kinetic exosite mutagenesis; domain deletion mutants tested across 20 vertebrate species for D4-induced activation and shear-dependent cleavage","pmids":["31439947","30700419"],"confidence":"High","gaps":["Full-length structure in complex with VWF not available","Precise conformational pathway from closed to open state not resolved"]},{"year":2019,"claim":"Epistatic interaction between ADAMTS13 deficiency and complement factor H mutation synergistically producing thrombotic microangiopathy established that the ADAMTS13-VWF axis and complement pathway cooperate in TMA pathogenesis.","evidence":"Double-mutant Adamts13⁻/⁻ × cfh-R1206 mice with multiple hematologic and renal endpoints","pmids":["31409673"],"confidence":"High","gaps":["Molecular mechanism linking VWF multimer accumulation to complement activation not defined","Whether complement inhibition can rescue ADAMTS13-deficient TMA"]},{"year":2020,"claim":"Kinetic dissection of conformational activation by anti-spacer and anti-CUB1 antibodies showed that allosteric relief of autoinhibition enhances kcat (not Km), clarifying that the autoinhibited conformation specifically restricts active-site catalytic turnover rather than substrate access, and refined epitope mapping identified three immunogenic hotspots in the spacer domain across 138 iTTP patients.","evidence":"kcat/Km determination with VWF A2 fragment proteolysis and conformational ELISA; library of 11 spacer-domain hybrids tested against 138 patient plasmas","pmids":["32196558","33171004"],"confidence":"High","gaps":["Structural basis of kcat enhancement at atomic resolution unknown","Whether gain-of-function ADAMTS13 variants could evade autoantibodies therapeutically"]},{"year":null,"claim":"Key open questions include the full-length structure of ADAMTS13 in complex with unfolded VWF, the precise conformational pathway from autoinhibited to active states, the physiological significance of the putative disulfide reductase activity of the distal domains, and the mechanistic basis by which ADAMTS13's C-terminal TSP1 repeats modulate angiogenesis independently of VWF proteolysis.","evidence":"","pmids":[],"confidence":"Low","gaps":["No full-length ADAMTS13–VWF complex structure","Disulfide reductase activity of CUB domains not validated independently","Angiogenic activity via TSP1 domains observed by a single group"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[0,1,2,4,6,8]},{"term_id":"GO:0016787","term_label":"hydrolase activity","supporting_discovery_ids":[0,1,4,6]}],"localization":[{"term_id":"GO:0005576","term_label":"extracellular region","supporting_discovery_ids":[1,2,7,11,12]},{"term_id":"GO:0005886","term_label":"plasma membrane","supporting_discovery_ids":[11]}],"pathway":[{"term_id":"R-HSA-109582","term_label":"Hemostasis","supporting_discovery_ids":[1,5,13,14,15]},{"term_id":"R-HSA-392499","term_label":"Metabolism of proteins","supporting_discovery_ids":[2,3]},{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[20,23,24]},{"term_id":"R-HSA-1643685","term_label":"Disease","supporting_discovery_ids":[13,15,20]}],"complexes":[],"partners":["VWF","POFUT2","FURIN","CFH","VEGFA"],"other_free_text":[]},"mechanistic_narrative":"ADAMTS13 is a plasma zinc metalloprotease that regulates hemostasis and vascular integrity by cleaving von Willebrand factor (VWF) at the Tyr1605-Met1606 bond within the A2 domain, thereby limiting the size and thrombogenic potential of ultra-large VWF multimers under fluid shear stress [PMID:11557746, PMID:12393399, PMID:21715306]. The enzyme adopts a closed, autoinhibited conformation in which the distal T8-CUB2 domains fold back onto the proximal MDTCS catalytic module; binding of the VWF D4 domain to the distal domains relieves this autoinhibition and allosterically activates the metalloprotease active site, so that VWF serves as both the activating cofactor and the substrate [PMID:25512528, PMID:31439947, PMID:32196558]. Efficient secretion of ADAMTS13 from hepatic and endothelial cells requires O-fucosylation of its thrombospondin type 1 repeats by POFUT2, and additional functionally active pools exist in platelets and glomerular endothelium [PMID:17395589, PMID:16176307, PMID:21720563]. Hereditary deficiency or acquired inhibitory autoantibodies—predominantly IgG4 targeting the spacer domain (Arg660/Tyr661/Tyr665 epitope region)—cause accumulation of ultra-large VWF multimers and thrombotic thrombocytopenic purpura, while ADAMTS13 deficiency also exacerbates ischemic stroke, atherosclerosis, and thrombotic microangiopathy through VWF-dependent mechanisms [PMID:23233642, PMID:19965676, PMID:24261607, PMID:31409673]."},"prefetch_data":{"uniprot":{"accession":"Q76LX8","full_name":"A disintegrin and metalloproteinase with thrombospondin motifs 13","aliases":["von Willebrand factor-cleaving protease","vWF-CP","vWF-cleaving protease"],"length_aa":1427,"mass_kda":153.6,"function":"Cleaves the vWF multimers in plasma into smaller forms thereby controlling vWF-mediated platelet thrombus formation","subcellular_location":"Secreted","url":"https://www.uniprot.org/uniprotkb/Q76LX8/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/ADAMTS13","classification":"Not Classified","n_dependent_lines":2,"n_total_lines":1208,"dependency_fraction":0.0016556291390728477},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/ADAMTS13","total_profiled":1310},"omim":[{"mim_id":"613554","title":"VON WILLEBRAND DISEASE, TYPE 2; 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apoptosis.","date":"2013","source":"Carcinogenesis","url":"https://pubmed.ncbi.nlm.nih.gov/23288922","citation_count":27,"is_preprint":false},{"pmid":"22433566","id":"PMC_22433566","title":"EGF activates TTP expression by activation of ELK-1 and EGR-1 transcription factors.","date":"2012","source":"BMC molecular biology","url":"https://pubmed.ncbi.nlm.nih.gov/22433566","citation_count":26,"is_preprint":false},{"pmid":"34505883","id":"PMC_34505883","title":"Plasma and rhADAMTS13 reduce trauma-induced organ failure by restoring the ADAMTS13-VWF axis.","date":"2021","source":"Blood advances","url":"https://pubmed.ncbi.nlm.nih.gov/34505883","citation_count":26,"is_preprint":false},{"pmid":"26542631","id":"PMC_26542631","title":"Increased expressions of ADAMTS-13 and apoptosis contribute to neuropathology during Toxoplasma gondii encephalitis in mice.","date":"2015","source":"Neuropathology : official journal of the Japanese Society of Neuropathology","url":"https://pubmed.ncbi.nlm.nih.gov/26542631","citation_count":25,"is_preprint":false},{"pmid":"32039742","id":"PMC_32039742","title":"A post-transcriptional program of chemoresistance by AU-rich elements and TTP in quiescent leukemic cells.","date":"2020","source":"Genome biology","url":"https://pubmed.ncbi.nlm.nih.gov/32039742","citation_count":25,"is_preprint":false},{"pmid":"24237985","id":"PMC_24237985","title":"HUS and TTP in Children.","date":"2013","source":"Pediatric clinics of North America","url":"https://pubmed.ncbi.nlm.nih.gov/24237985","citation_count":24,"is_preprint":false},{"pmid":"27771173","id":"PMC_27771173","title":"Complement activation, inflammation and relative ADAMTS13 deficiency in secondary thrombotic microangiopathies.","date":"2016","source":"Immunobiology","url":"https://pubmed.ncbi.nlm.nih.gov/27771173","citation_count":24,"is_preprint":false},{"pmid":"15662617","id":"PMC_15662617","title":"Interplay between ADAMTS13 and von Willebrand factor in inherited and acquired thrombotic microangiopathies.","date":"2005","source":"Seminars in hematology","url":"https://pubmed.ncbi.nlm.nih.gov/15662617","citation_count":23,"is_preprint":false},{"pmid":"31717307","id":"PMC_31717307","title":"The mRNA-binding Protein TTP/ZFP36 in Hepatocarcinogenesis and Hepatocellular Carcinoma.","date":"2019","source":"Cancers","url":"https://pubmed.ncbi.nlm.nih.gov/31717307","citation_count":22,"is_preprint":false},{"pmid":"35327035","id":"PMC_35327035","title":"The Intriguing Connections between von Willebrand Factor, ADAMTS13 and Cancer.","date":"2022","source":"Healthcare (Basel, Switzerland)","url":"https://pubmed.ncbi.nlm.nih.gov/35327035","citation_count":21,"is_preprint":false},{"pmid":"21720563","id":"PMC_21720563","title":"Phenotypic expression of ADAMTS13 in glomerular endothelial cells.","date":"2011","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/21720563","citation_count":21,"is_preprint":false},{"pmid":"18502798","id":"PMC_18502798","title":"Mechanistic studies on ADAMTS13 catalysis.","date":"2008","source":"Biophysical journal","url":"https://pubmed.ncbi.nlm.nih.gov/18502798","citation_count":21,"is_preprint":false},{"pmid":"33334760","id":"PMC_33334760","title":"COVID-19 presenting as thrombotic thrombocytopenic purpura (TTP).","date":"2020","source":"BMJ case reports","url":"https://pubmed.ncbi.nlm.nih.gov/33334760","citation_count":20,"is_preprint":false},{"pmid":"16113782","id":"PMC_16113782","title":"Transcriptional regulation of ADAMTS13.","date":"2005","source":"Thrombosis and haemostasis","url":"https://pubmed.ncbi.nlm.nih.gov/16113782","citation_count":20,"is_preprint":false},{"pmid":"33171004","id":"PMC_33171004","title":"Immunogenic hotspots in the spacer domain of ADAMTS13 in immune-mediated thrombotic thrombocytopenic purpura.","date":"2020","source":"Journal of thrombosis and haemostasis : JTH","url":"https://pubmed.ncbi.nlm.nih.gov/33171004","citation_count":20,"is_preprint":false},{"pmid":"31767150","id":"PMC_31767150","title":"Doxorubicin inhibits PD-L1 expression by enhancing TTP-mediated decay of PD-L1 mRNA in cancer cells.","date":"2019","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/31767150","citation_count":20,"is_preprint":false},{"pmid":"29777164","id":"PMC_29777164","title":"mRNA treatment produces sustained expression of enzymatically active human ADAMTS13 in mice.","date":"2018","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/29777164","citation_count":19,"is_preprint":false},{"pmid":"18536977","id":"PMC_18536977","title":"Tristetraprolin (TTP) gene polymorphisms in patients with rheumatoid arthritis and healthy individuals.","date":"2008","source":"Modern rheumatology","url":"https://pubmed.ncbi.nlm.nih.gov/18536977","citation_count":18,"is_preprint":false},{"pmid":"29203639","id":"PMC_29203639","title":"A Knock-In Tristetraprolin (TTP) Zinc Finger Point Mutation in Mice: Comparison with Complete TTP Deficiency.","date":"2018","source":"Molecular and cellular biology","url":"https://pubmed.ncbi.nlm.nih.gov/29203639","citation_count":18,"is_preprint":false},{"pmid":"35443560","id":"PMC_35443560","title":"Evidence of protective effects of recombinant ADAMTS13 in a humanized model of sickle cell disease.","date":"2022","source":"Haematologica","url":"https://pubmed.ncbi.nlm.nih.gov/35443560","citation_count":18,"is_preprint":false},{"pmid":"24106608","id":"PMC_24106608","title":"Proteolytic processing of von Willebrand factor by adamts13 and leukocyte proteases.","date":"2013","source":"Mediterranean journal of hematology and infectious diseases","url":"https://pubmed.ncbi.nlm.nih.gov/24106608","citation_count":17,"is_preprint":false},{"pmid":"35525391","id":"PMC_35525391","title":"Clinical implications of tristetraprolin (TTP) modulation in the treatment of inflammatory diseases.","date":"2022","source":"Pharmacology & therapeutics","url":"https://pubmed.ncbi.nlm.nih.gov/35525391","citation_count":16,"is_preprint":false},{"pmid":"29723192","id":"PMC_29723192","title":"The ARE-binding protein Tristetraprolin (TTP) is a novel target and mediator of calcineurin tumor suppressing function in the skin.","date":"2018","source":"PLoS genetics","url":"https://pubmed.ncbi.nlm.nih.gov/29723192","citation_count":16,"is_preprint":false},{"pmid":"11577265","id":"PMC_11577265","title":"Thrombotic thrombocytopenic purpura (TTP) and hemolytic uremic syndrome (HUS): the new thinking.","date":"2001","source":"Journal of thrombosis and thrombolysis","url":"https://pubmed.ncbi.nlm.nih.gov/11577265","citation_count":15,"is_preprint":false},{"pmid":"39125707","id":"PMC_39125707","title":"ADAMTS13 in the New Era of TTP.","date":"2024","source":"International journal of molecular sciences","url":"https://pubmed.ncbi.nlm.nih.gov/39125707","citation_count":14,"is_preprint":false},{"pmid":"28724967","id":"PMC_28724967","title":"A balancing act: RNA binding protein HuR/TTP axis in endometriosis patients.","date":"2017","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/28724967","citation_count":14,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":49422,"output_tokens":6405,"usd":0.122171},"stage2":{"model":"claude-opus-4-6","input_tokens":10119,"output_tokens":3781,"usd":0.21768},"total_usd":0.339851,"stage1_batch_id":"msgbatch_01TqNM6p6FsoMBT2jzQTwKBr","stage2_batch_id":"msgbatch_01Ya473eco8sTHbvRymQaU8o","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2001,\n      \"finding\": \"ADAMTS13 (VWF-cleaving protease) was identified as a new member of the ADAMTS family of metalloproteases with a domain architecture including: signal peptide, short propeptide (with furin consensus RQRR), reprolysin-like metalloprotease domain, disintegrin-like domain, thrombospondin-1 repeat, Cys-rich domain, ADAMTS spacer, seven additional thrombospondin-1 repeats, and two CUB domains. The Cys-rich domain contains an RGDS sequence. Sites for Zn2+ and Ca2+ are conserved in the protease domain.\",\n      \"method\": \"cDNA cloning, Northern blotting, sequence analysis\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — original cloning paper with full structural characterization, highly cited foundational study\",\n      \"pmids\": [\"11557746\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"Recombinant ADAMTS13 expressed in HEK 293 cells degrades VWF multimers and proteolytically cleaves VWF to the same fragments as plasma VWF-cleaving protease, confirming that ADAMTS13 is responsible for physiological proteolytic degradation of VWF multimers. Recombinant ADAMTS13-mediated degradation was entirely inhibited by plasma from a patient with acquired TTP.\",\n      \"method\": \"Expression of recombinant ADAMTS13 in HEK 293 cells, VWF multimer degradation assay, inhibition by patient plasma\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — direct functional reconstitution of enzymatic activity with controls\",\n      \"pmids\": [\"12393399\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"The ADAMTS13 propeptide is not required for folding, secretion, or enzymatic activity. Furin is the protease responsible for propeptide cleavage. Secreted pro-ADAMTS13 (with intact propeptide) has normal proteolytic activity toward VWF. In cells co-expressing ADAMTS13 and VWF, pro-ADAMTS13 can cleave pro-VWF intracellularly.\",\n      \"method\": \"Propeptide deletion and furin consensus site mutagenesis, expression in HeLa and furin-deficient LoVo cells, VWF cleavage assay\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — mutagenesis plus reconstitution in multiple cell lines with rigorous controls\",\n      \"pmids\": [\"12975358\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2007,\n      \"finding\": \"ADAMTS13 is O-fucosylated on at least six of its eight thrombospondin type 1 repeats (TSRs) with a glucose-β1,3-fucose disaccharide. This modification is required for efficient secretion of ADAMTS13; mutation of O-fucosylation sites or knockdown of POFUT2 (the transferase responsible) markedly reduces ADAMTS13 secretion.\",\n      \"method\": \"Mass spectral analysis of tryptic peptides, metabolic labeling with [3H]fucose, site-directed mutagenesis of modified serines, siRNA knockdown of POFUT2, GDP-fucose-deficient cell line\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — multiple orthogonal methods including MS, mutagenesis, and genetic depletion\",\n      \"pmids\": [\"17395589\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2010,\n      \"finding\": \"The ADAMTS13 metalloprotease domain contains two subsites critical for VWF cleavage: residues D187-R193 (VR1) are important for cleavage efficiency (kcat/Km reduced 2–10 fold by point mutations D187A, R190A, R193A), and residues D252-P256 (VR3) influence P1' amino acid specificity, shaping the S1' substrate-binding pocket.\",\n      \"method\": \"Composite sequence swaps and single-point mutagenesis in variable regions; kinetic analysis using VWF115 substrate and P1'-mutated VWF substrates\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — systematic mutagenesis with kinetic readout identifies catalytic subsites\",\n      \"pmids\": [\"20647566\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"ADAMTS13 proteolysis of VWF is controlled not by inhibitors but by conformational changes in VWF induced by elevated rheological shear forces, which unfold the VWF A2 domain to reveal cryptic exosites and the scissile bond (Tyr1605-Met1606). ADAMTS13 makes multiple exosite interactions with the unfolded A2 domain to position itself for cleavage.\",\n      \"method\": \"Review synthesizing biochemical and biophysical studies; supported by experimental literature on shear-dependent VWF unfolding and ADAMTS13 exosite mapping\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — synthesis of multiple experimental studies with strong mechanistic consensus\",\n      \"pmids\": [\"21715306\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2008,\n      \"finding\": \"ADAMTS13 catalysis of VWF A2-domain peptide (FRETS-VWF73) cleavage at Tyr1605-Met1606 is governed by two ionizable groups (pKa ~6.4 and ~4.0). The transition state is more ordered than the ground state (negative activation entropy). Residues Asp1653, Glu1655, Glu1660, Asp1663, and Thr1656 in the VWF A2 sequence form a 'hot spot' driving molecular recognition and allosteric regulation of ADAMTS13 binding. The product peptide (Met1606-Arg1668) acts as a hyperbolic mixed-type inhibitor.\",\n      \"method\": \"Fluorescence quenching (FRETS) kinetic assay across temperature and pH ranges; Co2+-substituted enzyme; product inhibition analysis; molecular modeling\",\n      \"journal\": \"Biophysical journal\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — detailed in vitro enzymology with multiple physical-chemical approaches\",\n      \"pmids\": [\"18502798\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"ADAMTS13 is allosterically regulated: the distal T8-CUB2 domains interact with and inhibit the proximal MDTCS catalytic domains. Binding of VWF D4 domain (or monoclonal antibodies) to the distal domains relieves this autoinhibition and markedly enhances substrate cleavage. Small angle X-ray scattering data support interdomain contact between distal T-CUB and proximal MDTCS domains in the closed, autoinhibited conformation.\",\n      \"method\": \"VWF D4-CUB binding assay, monoclonal antibody activation, kinetic proteolysis assays with truncation variants, small angle X-ray scattering (SAXS)\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — reconstitution with multiple domain variants, SAXS structural data, replicated with antibody activation\",\n      \"pmids\": [\"25512528\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"Crystal structure of ADAMTS13 (metalloprotease to spacer domains) reveals a latent conformation in which the active-site cleft is occluded, requiring allosteric change for substrate accommodation. Kinetic analyses show that cysteine-rich and spacer domain exosites bring enzyme and substrate into proximity, then disintegrin-like domain exosite binding to VWF allosterically activates the adjacent metalloprotease domain. VWF functions as both the activating cofactor and substrate.\",\n      \"method\": \"Crystal structure determination, kinetic analyses of VWF proteolysis by exosite mutants\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — crystal structure combined with kinetic functional validation, multiple orthogonal approaches\",\n      \"pmids\": [\"31439947\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"ADAMTS13 allosteric activation is conserved across vertebrates. The T7 and T8 distal thrombospondin domains are essential for allosteric activation by VWF D4 domain; deletion of T7 or T8 abolishes allosteric activation. Domains T3-T6 are dispensable for allosteric regulation and shear-dependent cleavage.\",\n      \"method\": \"Assay of ADAMTS13 from 20 placental mammals, birds, and amphibians; phylogenetic analysis; domain deletion mutants tested for VWF D4-induced activation and shear-dependent cleavage\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — systematic deletion mutagenesis combined with cross-species analysis, multiple orthogonal assays\",\n      \"pmids\": [\"30700419\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"Conformational activation of ADAMTS13 by anti-Spacer (3E4) or anti-CUB1 (17G2) monoclonal antibodies enhances ADAMTS13 catalytic efficiency (kcat/Km) ~2-fold. The mechanism is allosteric enhancement of metalloprotease domain active site function (increased kcat), not exposure of Spacer or cysteine-rich exosites or improved substrate binding (Km unchanged). A cryptic metalloprotease domain epitope is exposed upon mAb-induced conformational extension.\",\n      \"method\": \"Kinetic analyses of VWF A2 domain fragment proteolysis; ELISA-based conformational assay; kcat and Km determination\",\n      \"journal\": \"Blood advances\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 — kinetic dissection of activation mechanism with two independent mAbs and multiple readouts\",\n      \"pmids\": [\"32196558\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"Platelets contain functionally active ADAMTS13. Platelet lysates cleave endothelial cell-derived ultra-large VWF under static and flow conditions in a divalent cation-dependent manner; cleavage is inhibited by EDTA and by anti-ADAMTS13 antibody from a TTP patient. ADAMTS13 surface expression on platelets increases upon activation by thrombin receptor-activating peptide but not ADP.\",\n      \"method\": \"VWF cleavage assays with platelet lysates, antibody inhibition, flow cytometry, immunoblotting with four domain-specific antibodies\",\n      \"journal\": \"Journal of thrombosis and haemostasis : JTH\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — multiple antibodies, functional assay under flow, specific inhibition controls\",\n      \"pmids\": [\"16176307\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"Glomerular endothelial cells express and secrete functionally active ADAMTS13, which cleaves VWF. ADAMTS13 deficiency in mice results in thickening of glomerular capillaries with platelet deposition on the vessel wall.\",\n      \"method\": \"Real-time PCR, flow cytometry, immunofluorescence, immunoblotting, VWF multimer analysis in human glomerular endothelial cells; immunohistochemistry and electron microscopy in wild-type vs. Adamts13−/− mice\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — multiple orthogonal methods in both human cells and genetic mouse model with functional readout\",\n      \"pmids\": [\"21720563\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"ADAMTS13 deficiency aggravates ischemic brain damage in mice via the VWF-platelet axis: after reperfusion, Adamts13−/− mice show markedly decreased regional cerebral blood flow and larger infarct volumes compared to wild-type mice, demonstrating that ADAMTS13 protects against ischemia-reperfusion injury by regulating VWF-platelet interactions.\",\n      \"method\": \"Middle cerebral artery occlusion model in Adamts13−/− vs. wild-type mice; cerebral blood flow measurement; infarct volume quantification\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — genetic knockout model with defined mechanistic phenotype (VWF-platelet axis) and quantitative endpoints\",\n      \"pmids\": [\"19965676\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"The N-terminal MDTCS portion of ADAMTS13 is necessary and sufficient for proteolytic cleavage of VWF and attenuation of arterial/venous thrombosis. The distal portion (TSP1 2-8 repeats and CUB domains) may function as a disulfide bond reductase to prevent elongation of ultra-large VWF strings on activated endothelial cells. Proteolytic cleavage of VWF by ADAMTS13 is accelerated by FVIII and platelets under fluid shear stress; disruption of FVIII (or platelet glycoprotein 1bα)–VWF interactions dramatically impairs ADAMTS13-dependent proteolysis in vitro and in vivo.\",\n      \"method\": \"Domain truncation variants, thrombosis models in Adamts13−/− mice, in vitro VWF cleavage assays under flow\",\n      \"journal\": \"Journal of thrombosis and haemostasis : JTH\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — structure-function with multiple domain variants and genetic/in vitro validation\",\n      \"pmids\": [\"23809107\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"ADAMTS13 modulates atherosclerotic plaque progression through a VWF-dependent mechanism: deficiency of VWF in Adamts13−/−/ApoE−/− triple knockout mice completely reverses exacerbated atherosclerosis, macrophage/neutrophil infiltration, and interstitial collagen content compared with Adamts13−/−/ApoE−/− mice.\",\n      \"method\": \"Genetic epistasis using Adamts13−/−/Vwf−/−/ApoE−/− triple knockout mice on high-fat diet; aortic sinus cross-section analysis; immunohistochemistry\",\n      \"journal\": \"Journal of thrombosis and haemostasis : JTH\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — genetic epistasis with triple knockout cleanly placing ADAMTS13 upstream of VWF in atherosclerosis\",\n      \"pmids\": [\"24261607\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"Recombinant full-length ADAMTS13 promotes endothelial tube formation, proliferation, and migration in a dose-dependent manner independent of VEGF. ADAMTS13 also inhibits VEGF-induced angiogenesis, and these effects are mediated through the C-terminal TSP1 domains. Co-immunoprecipitation shows ADAMTS13 binds to VEGF via its TSP1 domain.\",\n      \"method\": \"Matrigel tube formation assay, proliferation counting assay, Boyden chamber migration assay, antibody blockade, truncation variants, co-immunoprecipitation\",\n      \"journal\": \"Microvascular research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — multiple functional assays with domain mapping and co-IP, but single lab\",\n      \"pmids\": [\"22626948\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"ADAMTS13 promotes angiogenesis via upregulation of VEGF expression and VEGFR2 phosphorylation in endothelial cells. The C-terminal TSP1 repeats (domains 2-8) are responsible for this angiogenic activity, as shown by structure-function analysis with truncation variants. Anti-VEGF IgG abrogates TSP1-domain-mediated effects on proliferation, migration, and VEGFR2 phosphorylation.\",\n      \"method\": \"Western blot, ELISA, VEGFR2 phosphorylation assay, proliferation and chemotaxis assays with TSP1-domain truncation variants and anti-VEGF antibody\",\n      \"journal\": \"Cellular and molecular life sciences : CMLS\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — multiple assays with domain mapping and antibody inhibition, single lab\",\n      \"pmids\": [\"24950743\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2009,\n      \"finding\": \"Autologous transplantation of hematopoietic progenitor cells transduced with lentiviral vector encoding ADAMTS13 restores plasma ADAMTS13 activity in Adamts13−/− mice, reduces VWF multimer size, and significantly prolongs ferric chloride-induced carotid arterial occlusion time, establishing hematopoietic cells as a source of functional ADAMTS13.\",\n      \"method\": \"Lentiviral transduction of hematopoietic progenitor cells, autologous transplantation in Adamts13−/− mice, plasma ADAMTS13 activity assay, VWF multimer analysis, ferric chloride thrombosis model\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — genetic correction with functional endpoint in vivo, multiple readouts\",\n      \"pmids\": [\"19141866\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2017,\n      \"finding\": \"ADAMTS13 controls vascular remodeling after ischemic stroke by modifying VWF reactivity. ADAMTS13-deficient mice show reduced neovascularization, impaired capillary perfusion, pericyte loss, and accelerated blood-brain barrier breakdown after stroke. VWF deficiency or anti-VWF antibody treatment reversed these deficits in Adamts13−/− mice, placing ADAMTS13 upstream of VWF in post-stroke angiogenesis.\",\n      \"method\": \"Adamts13−/−, Vwf−/−, and double-knockout mice in stroke model; microvascular analysis; angiopoietin-2 and galectin-3 measurement; adenoviral and recombinant protein rescue experiments; VEGFR2 antagonist\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — genetic epistasis with VWF knockout plus multiple rescue experiments establishing pathway\",\n      \"pmids\": [\"28428179\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"ADAMTS13 deficiency combined with complement factor H (cfh) heterozygous mutation (cfh R1206) synergistically induces thrombotic microangiopathy in mice, whereas either single defect alone does not cause overt disease, demonstrating epistatic interaction between the ADAMTS13-VWF axis and complement activation in TMA pathogenesis.\",\n      \"method\": \"Genetic epistasis using Adamts13−/−, cfh-mutant, and double-mutant mice; platelet counts, haptoglobin, erythrocyte fragmentation, LDH, BUN, creatinine, histology, immunohistochemistry\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — clean genetic epistasis with quantitative phenotypic endpoints across multiple genotypes\",\n      \"pmids\": [\"31409673\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"The transcriptional activity of the ADAMTS13 gene is not significantly altered by proinflammatory stimuli (endotoxin, TNF-α, IL-6, IL-1β), immunosuppressive agents (cyclosporine A, steroids, doxycycline), suggesting that diminished ADAMTS13 activity observed under various clinical conditions is not primarily due to transcriptional regulation.\",\n      \"method\": \"RNA analysis in liver homogenates and Hep3B liver cell culture; stimulation with multiple agents; in silico promoter analysis\",\n      \"journal\": \"Thrombosis and haemostasis\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — systematic stimulation panel with gene expression readout in relevant liver cell system\",\n      \"pmids\": [\"16113782\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"Platelet-delivered recombinant ADAMTS13 (expressed in transgenic platelets) is released upon stimulation with thrombin and collagen, inhibits arterial thrombosis after vascular injury, and prevents TTP syndrome in mice even in the absence of plasma ADAMTS13 activity or in the presence of anti-ADAMTS13 autoantibodies.\",\n      \"method\": \"Transgenic mice expressing ADAMTS13 in platelets, Western blot, FRET activity assay, ferric chloride thrombosis model, Shigatoxin-2 and VWF-induced TTP models\",\n      \"journal\": \"Blood\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — proof-of-concept in multiple in vivo models with functional activity assays\",\n      \"pmids\": [\"25800050\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"The epitope targeted by inhibitory anti-ADAMTS13 autoantibodies in acquired TTP is primarily within the spacer domain, specifically a surface comprising Arg660, Tyr661, and Tyr665, which contributes to productive binding of ADAMTS13 to unfolded VWF. IgG autoantibodies are predominantly of the IgG4 subclass.\",\n      \"method\": \"Epitope mapping, patient autoantibody characterization, domain-specific inhibition studies\",\n      \"journal\": \"Hematology. American Society of Hematology. Education Program\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — epitope mapping with functional inhibition data, supported across multiple patient studies\",\n      \"pmids\": [\"23233642\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"The immunogenic hotspots in the ADAMTS13 spacer domain targeted by anti-spacer autoantibodies in iTTP are amino acid regions 588-592, 602-610, and 657-666, identified using a library of 11 full-length ADAMTS13 spacer hybrids with corresponding ADAMTS1 spacer regions substituted.\",\n      \"method\": \"Library of 11 spacer hybrid ADAMTS13 proteins; ELISA epitope mapping with 138 iTTP patient plasma samples\",\n      \"journal\": \"Journal of thrombosis and haemostasis : JTH\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — systematic epitope mapping with large patient cohort and multiple hybrid constructs\",\n      \"pmids\": [\"33171004\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"ADAMTS13 is a constitutively active plasma metalloprotease that cleaves the Tyr1605-Met1606 bond in the VWF A2 domain under fluid shear stress; its activity is allosterically regulated by autoinhibitory contacts between C-terminal T8-CUB2 domains and the proximal MDTCS catalytic module, which are relieved by VWF D4 domain (or platelet/FVIII cofactor) binding to expose the active site, while the protease is secreted from liver and endothelial cells in a process requiring O-fucosylation of TSR domains, and its deficiency—whether hereditary or due to inhibitory autoantibodies targeting the spacer domain—causes accumulation of ultra-large VWF multimers and thrombotic thrombocytopenic purpura.\"\n}\n```","stage2_raw":"```json\n{\n  \"mechanistic_narrative\": \"ADAMTS13 is a plasma zinc metalloprotease that regulates hemostasis and vascular integrity by cleaving von Willebrand factor (VWF) at the Tyr1605-Met1606 bond within the A2 domain, thereby limiting the size and thrombogenic potential of ultra-large VWF multimers under fluid shear stress [PMID:11557746, PMID:12393399, PMID:21715306]. The enzyme adopts a closed, autoinhibited conformation in which the distal T8-CUB2 domains fold back onto the proximal MDTCS catalytic module; binding of the VWF D4 domain to the distal domains relieves this autoinhibition and allosterically activates the metalloprotease active site, so that VWF serves as both the activating cofactor and the substrate [PMID:25512528, PMID:31439947, PMID:32196558]. Efficient secretion of ADAMTS13 from hepatic and endothelial cells requires O-fucosylation of its thrombospondin type 1 repeats by POFUT2, and additional functionally active pools exist in platelets and glomerular endothelium [PMID:17395589, PMID:16176307, PMID:21720563]. Hereditary deficiency or acquired inhibitory autoantibodies—predominantly IgG4 targeting the spacer domain (Arg660/Tyr661/Tyr665 epitope region)—cause accumulation of ultra-large VWF multimers and thrombotic thrombocytopenic purpura, while ADAMTS13 deficiency also exacerbates ischemic stroke, atherosclerosis, and thrombotic microangiopathy through VWF-dependent mechanisms [PMID:23233642, PMID:19965676, PMID:24261607, PMID:31409673].\",\n  \"teleology\": [\n    {\n      \"year\": 2001,\n      \"claim\": \"Identification of the VWF-cleaving protease as a new ADAMTS family metalloprotease established its domain architecture—metalloprotease, disintegrin-like, TSR, Cys-rich, spacer, seven additional TSRs, and two CUB domains—providing the structural framework for all subsequent functional dissection.\",\n      \"evidence\": \"cDNA cloning, Northern blotting, and sequence analysis of the human gene\",\n      \"pmids\": [\"11557746\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"No three-dimensional structure yet available\", \"Expression sites and regulation unknown\", \"Catalytic mechanism not characterized\"]\n    },\n    {\n      \"year\": 2002,\n      \"claim\": \"Recombinant ADAMTS13 reconstituted VWF multimer degradation identical to the plasma protease and was inhibited by acquired TTP patient plasma, formally proving that ADAMTS13 is the physiological VWF-cleaving protease and that autoantibodies cause its functional deficiency.\",\n      \"evidence\": \"Recombinant expression in HEK 293 cells with VWF multimer degradation assay and patient-plasma inhibition\",\n      \"pmids\": [\"12393399\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Cleavage site specificity and kinetics not defined\", \"Propeptide role unknown\", \"In vivo relevance not yet demonstrated in genetic models\"]\n    },\n    {\n      \"year\": 2003,\n      \"claim\": \"Demonstrating that the propeptide is dispensable for folding, secretion, and enzymatic activity—and that pro-ADAMTS13 can cleave pro-VWF intracellularly—established that ADAMTS13 is constitutively active, unlike most ADAMTS family members that require propeptide removal for activation.\",\n      \"evidence\": \"Propeptide deletion and furin-site mutagenesis in HeLa and furin-deficient LoVo cells with VWF cleavage assays\",\n      \"pmids\": [\"12975358\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether intracellular cleavage is physiologically significant in vivo\", \"Regulatory mechanisms controlling activity remain unknown\"]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Discovery of functionally active ADAMTS13 in platelets and on the platelet surface (upregulated by thrombin) revealed a local, on-demand source of VWF-cleaving activity at sites of thrombus formation, beyond the known hepatic secretion.\",\n      \"evidence\": \"Platelet lysate VWF cleavage assay under static and flow conditions, flow cytometry, and inhibition by anti-ADAMTS13 antibody\",\n      \"pmids\": [\"16176307\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Relative contribution of platelet versus plasma ADAMTS13 in vivo unknown\", \"Mechanism of platelet ADAMTS13 storage and release not defined\"]\n    },\n    {\n      \"year\": 2007,\n      \"claim\": \"Identification of O-fucosylation on six of eight TSR domains and its requirement for secretion established a critical post-translational quality-control step: POFUT2-dependent glycosylation is necessary for ADAMTS13 to exit the ER efficiently.\",\n      \"evidence\": \"Mass spectrometry, metabolic labeling, site-directed mutagenesis of O-fucose sites, and siRNA knockdown of POFUT2\",\n      \"pmids\": [\"17395589\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether O-fucosylation also affects catalytic activity or stability in plasma\", \"Role of the glucose-β1,3-fucose disaccharide versus fucose alone\"]\n    },\n    {\n      \"year\": 2008,\n      \"claim\": \"Detailed enzymology of the Tyr1605-Met1606 cleavage reaction—revealing two catalytic ionizable groups, negative activation entropy, and a VWF A2 'hot spot' driving molecular recognition—provided the first kinetic and thermodynamic framework for ADAMTS13 catalysis.\",\n      \"evidence\": \"FRETS-VWF73 fluorescence quenching kinetics across pH and temperature; Co²⁺-substituted enzyme; product inhibition analysis\",\n      \"pmids\": [\"18502798\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural basis of the two ionizable groups not resolved\", \"Catalytic mechanism at atomic resolution still missing\"]\n    },\n    {\n      \"year\": 2009,\n      \"claim\": \"Genetic knockout studies in stroke and gene-therapy models demonstrated that ADAMTS13 deficiency worsens ischemia-reperfusion injury via the VWF-platelet axis and that hematopoietic cell-derived ADAMTS13 can rescue plasma activity, establishing both the in vivo protective role and a potential therapeutic source.\",\n      \"evidence\": \"Middle cerebral artery occlusion in Adamts13⁻/⁻ mice; lentiviral transduction of hematopoietic progenitor cells with autologous transplantation and thrombosis assays\",\n      \"pmids\": [\"19965676\", \"19141866\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Contribution of brain endothelial ADAMTS13 versus circulating protease not separated\", \"Optimal therapeutic delivery route undetermined\"]\n    },\n    {\n      \"year\": 2010,\n      \"claim\": \"Mutagenesis of the metalloprotease domain identified two substrate-contacting variable regions (VR1: D187-R193 for efficiency; VR3: D252-P256 for P1' specificity), beginning to define the molecular determinants of ADAMTS13's exquisite VWF selectivity.\",\n      \"evidence\": \"Composite sequence swaps and point mutations with kinetic analysis using VWF115 substrate\",\n      \"pmids\": [\"20647566\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structure of the enzyme–substrate complex not yet solved\", \"Full exosite map incomplete\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Synthesis of biochemical and biophysical evidence established that shear-induced unfolding of the VWF A2 domain—not a classical protease inhibitor—is the primary regulatory mechanism controlling ADAMTS13 proteolysis, and that glomerular endothelial cells are a functionally relevant local source of the enzyme.\",\n      \"evidence\": \"Review integrating shear-dependent VWF unfolding and exosite mapping studies; real-time PCR, immunofluorescence, and VWF multimer analysis in human glomerular endothelial cells and Adamts13⁻/⁻ mice\",\n      \"pmids\": [\"21715306\", \"21720563\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Quantitative threshold of shear force needed for A2 domain unfolding in vivo not precisely defined\", \"Relative importance of endothelial versus hepatic ADAMTS13 unclear\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Mapping the autoantibody epitope in acquired TTP to the spacer domain surface (Arg660/Tyr661/Tyr665) explained how autoantibodies inhibit VWF binding and established the spacer domain as both a critical substrate-recognition exosite and the principal immunogenic target.\",\n      \"evidence\": \"Epitope mapping with patient autoantibodies, domain-specific inhibition studies, IgG subclass characterization\",\n      \"pmids\": [\"23233642\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether epitope escape mutations can be engineered without loss of activity\", \"T-cell epitopes driving the autoimmune response not mapped\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Discovery of allosteric autoinhibition—where distal T8-CUB2 domains fold back on MDTCS and VWF D4 binding relieves this inhibition—fundamentally reframed ADAMTS13 as a conformationally regulated enzyme rather than a constitutively open protease, and genetic epistasis with VWF deficiency placed it upstream of VWF in atherosclerotic plaque progression.\",\n      \"evidence\": \"SAXS, domain truncation kinetics, and monoclonal antibody activation; Adamts13⁻/⁻/Vwf⁻/⁻/ApoE⁻/⁻ triple knockout mice on high-fat diet\",\n      \"pmids\": [\"25512528\", \"24261607\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Full-length structure in open and closed states not yet solved\", \"Whether additional cofactors modulate the open-closed transition in vivo\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"The crystal structure of the MDTCS module revealed a latent active-site conformation requiring allosteric opening, and cross-species analysis showed that T7-T8 domains are specifically essential for VWF D4-mediated allosteric activation, while T3-T6 are dispensable.\",\n      \"evidence\": \"Crystal structure determination with kinetic exosite mutagenesis; domain deletion mutants tested across 20 vertebrate species for D4-induced activation and shear-dependent cleavage\",\n      \"pmids\": [\"31439947\", \"30700419\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Full-length structure in complex with VWF not available\", \"Precise conformational pathway from closed to open state not resolved\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Epistatic interaction between ADAMTS13 deficiency and complement factor H mutation synergistically producing thrombotic microangiopathy established that the ADAMTS13-VWF axis and complement pathway cooperate in TMA pathogenesis.\",\n      \"evidence\": \"Double-mutant Adamts13⁻/⁻ × cfh-R1206 mice with multiple hematologic and renal endpoints\",\n      \"pmids\": [\"31409673\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular mechanism linking VWF multimer accumulation to complement activation not defined\", \"Whether complement inhibition can rescue ADAMTS13-deficient TMA\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Kinetic dissection of conformational activation by anti-spacer and anti-CUB1 antibodies showed that allosteric relief of autoinhibition enhances kcat (not Km), clarifying that the autoinhibited conformation specifically restricts active-site catalytic turnover rather than substrate access, and refined epitope mapping identified three immunogenic hotspots in the spacer domain across 138 iTTP patients.\",\n      \"evidence\": \"kcat/Km determination with VWF A2 fragment proteolysis and conformational ELISA; library of 11 spacer-domain hybrids tested against 138 patient plasmas\",\n      \"pmids\": [\"32196558\", \"33171004\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural basis of kcat enhancement at atomic resolution unknown\", \"Whether gain-of-function ADAMTS13 variants could evade autoantibodies therapeutically\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"Key open questions include the full-length structure of ADAMTS13 in complex with unfolded VWF, the precise conformational pathway from autoinhibited to active states, the physiological significance of the putative disulfide reductase activity of the distal domains, and the mechanistic basis by which ADAMTS13's C-terminal TSP1 repeats modulate angiogenesis independently of VWF proteolysis.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No full-length ADAMTS13–VWF complex structure\", \"Disulfide reductase activity of CUB domains not validated independently\", \"Angiogenic activity via TSP1 domains observed by a single group\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [0, 1, 2, 4, 6, 8]},\n      {\"term_id\": \"GO:0016787\", \"supporting_discovery_ids\": [0, 1, 4, 6]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005576\", \"supporting_discovery_ids\": [1, 2, 7, 11, 12]},\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [11]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-109582\", \"supporting_discovery_ids\": [1, 5, 13, 14, 15]},\n      {\"term_id\": \"R-HSA-392499\", \"supporting_discovery_ids\": [2, 3]},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [20, 23, 24]},\n      {\"term_id\": \"R-HSA-1643685\", \"supporting_discovery_ids\": [13, 15, 20]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\n      \"VWF\",\n      \"POFUT2\",\n      \"FURIN\",\n      \"CFH\",\n      \"VEGFA\"\n    ],\n    \"other_free_text\": []\n  }\n}\n```"}