{"gene":"PF3D7_1457500","run_date":"2026-09-08T14:19:07","timeline":{"discoveries":[{"year":2004,"finding":"Endogenous PfVps4 (PF3D7_1457500) localizes to the cytoplasm of P. falciparum trophozoites; an ATP-hydrolysis-blocking mutation causes it to relocalize from the cytosol to large vesicular structures, demonstrating that ATP hydrolysis is required for its normal cytosolic distribution.","method":"Immunofluorescence of endogenous protein; transient expression of ATPase-dead mutant constructs in P. falciparum, Toxoplasma gondii, and COS cells","journal":"Journal of cell science","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct localization by live/fixed imaging with two independent dominant-negative mutants (ATP-hydrolysis-blocked and ATP-binding-blocked), single lab","pmids":["15252121"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2004,"finding":"Expression of ATP-hydrolysis-defective or ATP-binding-defective PfVps4 (PF3D7_1457500) generates large cholesterol-enriched multivesicular bodies in heterologous systems, establishing that VPS4 function is required for multivesicular body formation in the parasite.","method":"Transient expression of dominant-negative PfVps4 mutants in COS cells and T. gondii; cholesterol staining and co-localization with early endosome markers","journal":"Journal of cell science","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — functional dominant-negative approach with two independent mutants and organelle markers, single lab, heterologous systems used as validation","pmids":["15252121"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2026,"finding":"Conditional knockdown of PfVps4 (PF3D7_1457500) in P. falciparum causes an 84% reduction in asexual progeny numbers and decreased merozoite numbers, establishing that PfVps4 ATPase activity is essential for intraerythrocytic proliferation.","method":"CRISPR-Cas9-mediated conditional knockdown; tightly synchronized cultures; Western blotting and immunofluorescence assays to confirm knockdown","journal":"Parasites & vectors","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — clean conditional KO with specific proliferative phenotype, single lab, single study","pmids":["41957628"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":"3D7"},{"species":"Plasmodium falciparum","strain":"NF54"}]},{"year":2026,"finding":"Conditional knockdown of PfVps4 (PF3D7_1457500) reduces gametocyte conversion rate by 46% without affecting subsequent gametocyte maturation, indicating PfVps4 is required for efficient sexual conversion but not for later gametocyte development.","method":"CRISPR-Cas9 conditional knockdown; gametocytogenesis assay with synchronized cultures; comparison of conversion rates","journal":"Parasites & vectors","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — conditional KO with stage-specific phenotypic readout, single lab, single study","pmids":["41957628"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":"3D7"},{"species":"Plasmodium falciparum","strain":"NF54"}]},{"year":2026,"finding":"In vitro biochemical characterization of purified PfVps4 (PF3D7_1457500) shows it is an AAA-ATPase with optimal activity at pH 7.5 and 37°C, Mg²⁺-dependent, with a Vmax of 2.23 ± 0.053 U/mg and Km of 0.086 mM; site-directed mutagenesis of canonical catalytic residues D213 and E214 abolishes activity, confirming their essential catalytic role.","method":"In vitro protein purification; enzyme kinetics assays; site-directed mutagenesis","journal":"Parasites & vectors","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro reconstitution with enzyme kinetics and mutagenesis of active-site residues, single lab","pmids":["41957628"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2026,"finding":"Species-specific residues T161 and I288 of PfVps4 (PF3D7_1457500) are required for enzymatic activity, as demonstrated by site-directed mutagenesis combined with enzyme kinetics.","method":"Site-directed mutagenesis; in vitro ATPase activity assays","journal":"Parasites & vectors","confidence":"Medium","confidence_rationale":"Tier 1 / Weak — in vitro mutagenesis with activity assay, single lab, single study","pmids":["41957628"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2026,"finding":"The cofactor PfVta1 significantly enhances PfVps4 (PF3D7_1457500) ATPase activity in vitro, establishing a functional activating interaction between these two proteins.","method":"In vitro ATPase activity assay with and without recombinant PfVta1; enzyme kinetics","journal":"Parasites & vectors","confidence":"Medium","confidence_rationale":"Tier 1 / Weak — biochemical reconstitution showing cofactor activation, single lab, single study","pmids":["41957628"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]}],"current_model":"PfVps4 (PF3D7_1457500) is a Mg²⁺-dependent AAA-ATPase whose ATP hydrolysis activity — dependent on catalytic residues D213/E214 and species-specific residues T161/I288, and stimulated by the cofactor PfVta1 — is required for cytosolic localization and multivesicular body biogenesis in P. falciparum trophozoites; conditional knockdown establishes it as essential for intraerythrocytic proliferation and efficient sexual conversion to gametocytes."},"narrative":{"mechanistic_narrative":"PF3D7_1457500 (VPS4) is a Mg²⁺-dependent AAA-ATPase of P. falciparum whose ATP hydrolysis activity drives multivesicular body biogenesis and is required for intraerythrocytic proliferation [PMID:15252121, PMID:41957628]. The endogenous protein resides in the cytoplasm of trophozoites, and blocking ATP hydrolysis relocalizes it from the cytosol onto large vesicular structures, demonstrating that its catalytic cycle governs its normal cytosolic distribution [PMID:15252121]. Expression of ATP-hydrolysis- or ATP-binding-defective VPS4 generates enlarged cholesterol-enriched multivesicular bodies, placing the enzyme at the core of parasite endosomal/MVB formation [PMID:15252121]. In vitro, purified VPS4 is a bona fide AAA-ATPase (optimal at pH 7.5/37°C, Mg²⁺-dependent) whose activity depends on the canonical catalytic residues D213 and E214 and on the species-specific residues T161 and I288, and is stimulated by the cofactor PfVta1 [PMID:41957628]. Conditional knockdown reduces asexual progeny by 84% and merozoite numbers, establishing essentiality for blood-stage proliferation, and additionally lowers gametocyte conversion by 46% without impairing subsequent maturation, implicating VPS4 in efficient sexual conversion [PMID:41957628]. Beyond these findings, no further binding partners or downstream substrates of PF3D7_1457500 have been characterized in the available corpus.","teleology":[{"year":2004,"claim":"To learn whether VPS4's ATPase cycle controls its subcellular behavior, its localization was probed in normal versus catalytically dead states, establishing that ATP hydrolysis maintains its cytosolic distribution.","evidence":"Immunofluorescence of endogenous protein plus transient expression of ATPase-dead and ATP-binding-dead mutants in P. falciparum, T. gondii, and COS cells","pmids":["15252121"],"confidence":"Medium","gaps":["Dominant-negative relocalization does not identify the membranes/vesicles VPS4 acts on directly","No in situ validation of MVB engagement in native parasite cytosol"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2004,"claim":"To connect VPS4 to an organellar process, dominant-negative mutants were tested for effects on membrane compartments, showing that VPS4 function is required for multivesicular body formation.","evidence":"Transient expression of dominant-negative mutants in COS cells and T. gondii with cholesterol staining and early-endosome marker co-localization","pmids":["15252121"],"confidence":"Medium","gaps":["MVB phenotype demonstrated largely in heterologous systems rather than natively in P. falciparum","No identification of cargo sorted through these MVBs"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2026,"claim":"To move from dominant-negative inference to loss-of-function, conditional knockdown tested whether VPS4 is needed for parasite growth, establishing it as essential for intraerythrocytic proliferation.","evidence":"CRISPR-Cas9 conditional knockdown in synchronized 3D7/NF54 cultures with Western blot and IFA confirmation","pmids":["41957628"],"confidence":"Medium","gaps":["Single lab, single study","Proliferation defect not resolved to a specific step in the asexual cycle beyond reduced merozoite numbers"],"attribution":[{"species":"Plasmodium falciparum","strain":"3D7"},{"species":"Plasmodium falciparum","strain":"NF54"}]},{"year":2026,"claim":"To test a role beyond asexual growth, knockdown was assessed across sexual commitment, showing VPS4 is required for efficient conversion to gametocytes but not for their later maturation.","evidence":"CRISPR-Cas9 conditional knockdown with gametocytogenesis conversion-rate assay","pmids":["41957628"],"confidence":"Medium","gaps":["Mechanism linking VPS4 ATPase activity to commitment signaling is unknown","Single study"],"attribution":[{"species":"Plasmodium falciparum","strain":"3D7"},{"species":"Plasmodium falciparum","strain":"NF54"}]},{"year":2026,"claim":"To define the enzyme biochemically, purified VPS4 was subjected to kinetics and active-site mutagenesis, confirming Mg²⁺-dependent AAA-ATPase activity requiring catalytic residues D213/E214.","evidence":"In vitro purification, enzyme kinetics (Vmax 2.23 U/mg, Km 0.086 mM), and site-directed mutagenesis of D213/E214","pmids":["41957628"],"confidence":"High","gaps":["No structural model of the ATPase or its oligomeric assembly","Physiological substrates/machinery acted upon not defined"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2026,"claim":"To probe parasite-specific features exploitable for selectivity, non-canonical residues were mutated, showing T161 and I288 are required for enzymatic activity.","evidence":"Site-directed mutagenesis with in vitro ATPase assays","pmids":["41957628"],"confidence":"Medium","gaps":["Structural basis for how T161/I288 support catalysis is not established","Single study"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2026,"claim":"To identify a functional partner regulating activity, the cofactor PfVta1 was tested, establishing it as an activator of VPS4 ATPase activity.","evidence":"In vitro ATPase kinetics with and without recombinant PfVta1","pmids":["41957628"],"confidence":"Medium","gaps":["Physical basis and stoichiometry of the VPS4–PfVta1 interaction not defined","In vivo dependence of VPS4 function on PfVta1 not tested"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":null,"claim":"It remains unknown what protein substrates VPS4 disassembles, how its ATPase cycle is structurally coupled to MVB membrane remodeling, and how these activities mechanistically drive both proliferation and sexual conversion.","evidence":"","pmids":[],"confidence":"Medium","gaps":["No structural model or oligomeric assembly defined","No identified substrate complex acted upon by VPS4","Mechanistic link between ATPase activity and the sexual-conversion phenotype unresolved"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]}],"mechanism_profile":{"molecular_activity":[],"localization":[],"pathway":[],"complexes":[],"partners":["PFVTA1"],"other_free_text":["Mg²⁺-dependent AAA-ATPase with catalytic residues D213/E214 and species-specific residues T161/I288 required for activity [#4, #5]","ATPase activity stimulated by cofactor PfVta1 [#6]","Essential for intraerythrocytic proliferation (84% reduction in asexual progeny on knockdown) and required for efficient gametocyte conversion [#2, #3]"]}},"prefetch_data":{},"corpus_meta":[{"pmid":"15252121","id":"PMC_15252121","title":"The Plasmodium falciparum Vps4 homolog mediates multivesicular body formation.","date":"2004","source":"Journal of cell science","url":"https://pubmed.ncbi.nlm.nih.gov/15252121","citation_count":39,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["VPS4","PfVPS4"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"41957628","id":"PMC_41957628","title":"PfVPS4, an ESCRT AAA-ATPase, is essential for asexual proliferation and gametocyte sexual conversion in Plasmodium falciparum.","date":"2026","source":"Parasites & vectors","url":"https://pubmed.ncbi.nlm.nih.gov/41957628","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["vacuolar protein sorting-associated protein 4","PfVPS4"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":4254,"output_tokens":1915,"usd":0.020744,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":7054,"output_tokens":3162,"usd":0.05716,"stage2_stop_reason":"end_turn"},"total_usd":0.077904,"stage1_batch_id":"msgbatch_011BPjN2BL33F8cJXD1r7V7T","stage2_batch_id":"msgbatch_01Qwg6vP7qxkpL6Gsc7tajsb","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2004,\n      \"finding\": \"Endogenous PfVps4 (PF3D7_1457500) localizes to the cytoplasm of P. falciparum trophozoites; an ATP-hydrolysis-blocking mutation causes it to relocalize from the cytosol to large vesicular structures, demonstrating that ATP hydrolysis is required for its normal cytosolic distribution.\",\n      \"method\": \"Immunofluorescence of endogenous protein; transient expression of ATPase-dead mutant constructs in P. falciparum, Toxoplasma gondii, and COS cells\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct localization by live/fixed imaging with two independent dominant-negative mutants (ATP-hydrolysis-blocked and ATP-binding-blocked), single lab\",\n      \"pmids\": [\"15252121\"],\n      \"is_preprint\": false,\n      \"attribution\": [\n        {\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}\n      ]\n    },\n    {\n      \"year\": 2004,\n      \"finding\": \"Expression of ATP-hydrolysis-defective or ATP-binding-defective PfVps4 (PF3D7_1457500) generates large cholesterol-enriched multivesicular bodies in heterologous systems, establishing that VPS4 function is required for multivesicular body formation in the parasite.\",\n      \"method\": \"Transient expression of dominant-negative PfVps4 mutants in COS cells and T. gondii; cholesterol staining and co-localization with early endosome markers\",\n      \"journal\": \"Journal of cell science\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — functional dominant-negative approach with two independent mutants and organelle markers, single lab, heterologous systems used as validation\",\n      \"pmids\": [\"15252121\"],\n      \"is_preprint\": false,\n      \"attribution\": [\n        {\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}\n      ]\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"Conditional knockdown of PfVps4 (PF3D7_1457500) in P. falciparum causes an 84% reduction in asexual progeny numbers and decreased merozoite numbers, establishing that PfVps4 ATPase activity is essential for intraerythrocytic proliferation.\",\n      \"method\": \"CRISPR-Cas9-mediated conditional knockdown; tightly synchronized cultures; Western blotting and immunofluorescence assays to confirm knockdown\",\n      \"journal\": \"Parasites & vectors\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — clean conditional KO with specific proliferative phenotype, single lab, single study\",\n      \"pmids\": [\"41957628\"],\n      \"is_preprint\": false,\n      \"attribution\": [\n        {\"species\": \"Plasmodium falciparum\", \"strain\": \"3D7\"},\n        {\"species\": \"Plasmodium falciparum\", \"strain\": \"NF54\"}\n      ]\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"Conditional knockdown of PfVps4 (PF3D7_1457500) reduces gametocyte conversion rate by 46% without affecting subsequent gametocyte maturation, indicating PfVps4 is required for efficient sexual conversion but not for later gametocyte development.\",\n      \"method\": \"CRISPR-Cas9 conditional knockdown; gametocytogenesis assay with synchronized cultures; comparison of conversion rates\",\n      \"journal\": \"Parasites & vectors\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — conditional KO with stage-specific phenotypic readout, single lab, single study\",\n      \"pmids\": [\"41957628\"],\n      \"is_preprint\": false,\n      \"attribution\": [\n        {\"species\": \"Plasmodium falciparum\", \"strain\": \"3D7\"},\n        {\"species\": \"Plasmodium falciparum\", \"strain\": \"NF54\"}\n      ]\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"In vitro biochemical characterization of purified PfVps4 (PF3D7_1457500) shows it is an AAA-ATPase with optimal activity at pH 7.5 and 37°C, Mg²⁺-dependent, with a Vmax of 2.23 ± 0.053 U/mg and Km of 0.086 mM; site-directed mutagenesis of canonical catalytic residues D213 and E214 abolishes activity, confirming their essential catalytic role.\",\n      \"method\": \"In vitro protein purification; enzyme kinetics assays; site-directed mutagenesis\",\n      \"journal\": \"Parasites & vectors\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro reconstitution with enzyme kinetics and mutagenesis of active-site residues, single lab\",\n      \"pmids\": [\"41957628\"],\n      \"is_preprint\": false,\n      \"attribution\": [\n        {\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}\n      ]\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"Species-specific residues T161 and I288 of PfVps4 (PF3D7_1457500) are required for enzymatic activity, as demonstrated by site-directed mutagenesis combined with enzyme kinetics.\",\n      \"method\": \"Site-directed mutagenesis; in vitro ATPase activity assays\",\n      \"journal\": \"Parasites & vectors\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Weak — in vitro mutagenesis with activity assay, single lab, single study\",\n      \"pmids\": [\"41957628\"],\n      \"is_preprint\": false,\n      \"attribution\": [\n        {\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}\n      ]\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"The cofactor PfVta1 significantly enhances PfVps4 (PF3D7_1457500) ATPase activity in vitro, establishing a functional activating interaction between these two proteins.\",\n      \"method\": \"In vitro ATPase activity assay with and without recombinant PfVta1; enzyme kinetics\",\n      \"journal\": \"Parasites & vectors\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Weak — biochemical reconstitution showing cofactor activation, single lab, single study\",\n      \"pmids\": [\"41957628\"],\n      \"is_preprint\": false,\n      \"attribution\": [\n        {\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}\n      ]\n    }\n  ],\n  \"current_model\": \"PfVps4 (PF3D7_1457500) is a Mg²⁺-dependent AAA-ATPase whose ATP hydrolysis activity — dependent on catalytic residues D213/E214 and species-specific residues T161/I288, and stimulated by the cofactor PfVta1 — is required for cytosolic localization and multivesicular body biogenesis in P. falciparum trophozoites; conditional knockdown establishes it as essential for intraerythrocytic proliferation and efficient sexual conversion to gametocytes.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"PF3D7_1457500 (VPS4) is a Mg²⁺-dependent AAA-ATPase of P. falciparum whose ATP hydrolysis activity drives multivesicular body biogenesis and is required for intraerythrocytic proliferation [#1, #2, #4]. The endogenous protein resides in the cytoplasm of trophozoites, and blocking ATP hydrolysis relocalizes it from the cytosol onto large vesicular structures, demonstrating that its catalytic cycle governs its normal cytosolic distribution [#0]. Expression of ATP-hydrolysis- or ATP-binding-defective VPS4 generates enlarged cholesterol-enriched multivesicular bodies, placing the enzyme at the core of parasite endosomal/MVB formation [#1]. In vitro, purified VPS4 is a bona fide AAA-ATPase (optimal at pH 7.5/37°C, Mg²⁺-dependent) whose activity depends on the canonical catalytic residues D213 and E214 and on the species-specific residues T161 and I288, and is stimulated by the cofactor PfVta1 [#4, #5, #6]. Conditional knockdown reduces asexual progeny by 84% and merozoite numbers, establishing essentiality for blood-stage proliferation, and additionally lowers gametocyte conversion by 46% without impairing subsequent maturation, implicating VPS4 in efficient sexual conversion [#2, #3]. Beyond these findings, no further binding partners or downstream substrates of PF3D7_1457500 have been characterized in the available corpus.\",\n  \"teleology\": [\n    {\n      \"year\": 2004,\n      \"claim\": \"To learn whether VPS4's ATPase cycle controls its subcellular behavior, its localization was probed in normal versus catalytically dead states, establishing that ATP hydrolysis maintains its cytosolic distribution.\",\n      \"evidence\": \"Immunofluorescence of endogenous protein plus transient expression of ATPase-dead and ATP-binding-dead mutants in P. falciparum, T. gondii, and COS cells\",\n      \"pmids\": [\"15252121\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Dominant-negative relocalization does not identify the membranes/vesicles VPS4 acts on directly\",\n        \"No in situ validation of MVB engagement in native parasite cytosol\"\n      ],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2004,\n      \"claim\": \"To connect VPS4 to an organellar process, dominant-negative mutants were tested for effects on membrane compartments, showing that VPS4 function is required for multivesicular body formation.\",\n      \"evidence\": \"Transient expression of dominant-negative mutants in COS cells and T. gondii with cholesterol staining and early-endosome marker co-localization\",\n      \"pmids\": [\"15252121\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"MVB phenotype demonstrated largely in heterologous systems rather than natively in P. falciparum\",\n        \"No identification of cargo sorted through these MVBs\"\n      ],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2026,\n      \"claim\": \"To move from dominant-negative inference to loss-of-function, conditional knockdown tested whether VPS4 is needed for parasite growth, establishing it as essential for intraerythrocytic proliferation.\",\n      \"evidence\": \"CRISPR-Cas9 conditional knockdown in synchronized 3D7/NF54 cultures with Western blot and IFA confirmation\",\n      \"pmids\": [\"41957628\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Single lab, single study\",\n        \"Proliferation defect not resolved to a specific step in the asexual cycle beyond reduced merozoite numbers\"\n      ],\n      \"attribution\": [\n        {\"species\": \"Plasmodium falciparum\", \"strain\": \"3D7\"},\n        {\"species\": \"Plasmodium falciparum\", \"strain\": \"NF54\"}\n      ]\n    },\n    {\n      \"year\": 2026,\n      \"claim\": \"To test a role beyond asexual growth, knockdown was assessed across sexual commitment, showing VPS4 is required for efficient conversion to gametocytes but not for their later maturation.\",\n      \"evidence\": \"CRISPR-Cas9 conditional knockdown with gametocytogenesis conversion-rate assay\",\n      \"pmids\": [\"41957628\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Mechanism linking VPS4 ATPase activity to commitment signaling is unknown\",\n        \"Single study\"\n      ],\n      \"attribution\": [\n        {\"species\": \"Plasmodium falciparum\", \"strain\": \"3D7\"},\n        {\"species\": \"Plasmodium falciparum\", \"strain\": \"NF54\"}\n      ]\n    },\n    {\n      \"year\": 2026,\n      \"claim\": \"To define the enzyme biochemically, purified VPS4 was subjected to kinetics and active-site mutagenesis, confirming Mg²⁺-dependent AAA-ATPase activity requiring catalytic residues D213/E214.\",\n      \"evidence\": \"In vitro purification, enzyme kinetics (Vmax 2.23 U/mg, Km 0.086 mM), and site-directed mutagenesis of D213/E214\",\n      \"pmids\": [\"41957628\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"No structural model of the ATPase or its oligomeric assembly\",\n        \"Physiological substrates/machinery acted upon not defined\"\n      ],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2026,\n      \"claim\": \"To probe parasite-specific features exploitable for selectivity, non-canonical residues were mutated, showing T161 and I288 are required for enzymatic activity.\",\n      \"evidence\": \"Site-directed mutagenesis with in vitro ATPase assays\",\n      \"pmids\": [\"41957628\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Structural basis for how T161/I288 support catalysis is not established\",\n        \"Single study\"\n      ],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2026,\n      \"claim\": \"To identify a functional partner regulating activity, the cofactor PfVta1 was tested, establishing it as an activator of VPS4 ATPase activity.\",\n      \"evidence\": \"In vitro ATPase kinetics with and without recombinant PfVta1\",\n      \"pmids\": [\"41957628\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"Physical basis and stoichiometry of the VPS4–PfVta1 interaction not defined\",\n        \"In vivo dependence of VPS4 function on PfVta1 not tested\"\n      ],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"It remains unknown what protein substrates VPS4 disassembles, how its ATPase cycle is structurally coupled to MVB membrane remodeling, and how these activities mechanistically drive both proliferation and sexual conversion.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\n        \"No structural model or oligomeric assembly defined\",\n        \"No identified substrate complex acted upon by VPS4\",\n        \"Mechanistic link between ATPase activity and the sexual-conversion phenotype unresolved\"\n      ],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0016887\", \"supporting_discovery_ids\": [4, 5]},\n      {\"term_id\": \"GO:0005524\", \"supporting_discovery_ids\": [0, 4]},\n      {\"term_id\": \"GO:0000287\", \"supporting_discovery_ids\": [4]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [0]},\n      {\"term_id\": \"GO:0005771\", \"supporting_discovery_ids\": [1]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"GO:0016050\", \"supporting_discovery_ids\": [1]},\n      {\"term_id\": \"GO:0060623\", \"supporting_discovery_ids\": [2, 3]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"PfVta1\"],\n    \"other_free_text\": [\n      \"Mg²⁺-dependent AAA-ATPase with catalytic residues D213/E214 and species-specific residues T161/I288 required for activity [#4, #5]\",\n      \"ATPase activity stimulated by cofactor PfVta1 [#6]\",\n      \"Essential for intraerythrocytic proliferation (84% reduction in asexual progeny on knockdown) and required for efficient gametocyte conversion [#2, #3]\"\n    ]\n  }\n}","audit_flag":null,"strains":[{"strain":"Pf3D7","gene_id":"PF3D7_1457500","name":null,"product":"vacuolar protein sorting-associated protein 4","uniprot_acc":null,"chromosome":null,"start_pos":null},{"strain":"PbANKA","gene_id":"PBANKA_1321200","name":null,"product":"vacuolar protein sorting-associated protein 4, putative","uniprot_acc":null,"chromosome":null,"start_pos":null},{"strain":"PkH","gene_id":"PKNH_1224300","name":null,"product":"vacuolar protein sorting-associated protein 4, putative","uniprot_acc":null,"chromosome":null,"start_pos":null}],"gene_symbol":"VPS4","evaluation":null}