{"gene":"PF3D7_1115300","run_date":"2026-09-08T12:51:28","timeline":{"discoveries":[{"year":2006,"finding":"Recombinant FP2B expressed in bacteria exhibits cysteine protease activity, demonstrated by cleavage of a fluorescent peptide substrate and in-gel gelatin zymography. FP2B is transcribed at the trophozoite stage of P. falciparum.","method":"Recombinant protein expression in bacteria, fluorescent peptide substrate cleavage assay, in-gel gelatin zymography, RT-PCR","journal":"Blood cells, molecules & diseases","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct enzymatic assay with two orthogonal methods (fluorescent substrate + zymography), single lab","pmids":["16595182"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2006,"finding":"Recombinant FP2B cleaves host erythrocyte ankyrin but not protein 4.1, as assessed by an erythrocyte inside-out-vesicle assay in vitro, suggesting a selective proteolytic role in parasite egress from red blood cells.","method":"Erythrocyte inside-out-vesicle assay with recombinant FP2B","journal":"Blood cells, molecules & diseases","confidence":"Medium","confidence_rationale":"Tier 2 / Weak — direct in vitro substrate cleavage assay, single lab, single method","pmids":["16595182"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2005,"finding":"The mature protease domain of FP2B (falcipain-2'), expressed in E. coli and refolded, is an active hemoglobinase with biochemical properties similar to falcipain-2: pH optima of 5.5–7.0, reducing agent requirement, and similar substrate preference. Both enzymes readily hydrolyze hemoglobin in vitro.","method":"Recombinant protein expression and refolding in E. coli, enzymatic activity assays with peptide substrates and hemoglobin, inhibitor profiling","journal":"Experimental parasitology","confidence":"Medium","confidence_rationale":"Tier 1 / Moderate — in vitro reconstitution of enzymatic activity with multiple substrate assays, single lab","pmids":["16337629"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2005,"finding":"FP2B (PF11_0161) is highly expressed as an active enzyme during the erythrocytic stage. Site-directed mutagenesis of active-site-proximate residues (S149A/I85M and S149A/A151D double mutants) rendered FP2B completely inactive, identifying I85, S149, and A151 as structurally critical residues. FP2B is more tolerant to substitution at position 149 than FP2A despite 97.5% sequence identity between the two isozymes.","method":"Active-site mutagenesis, enzymatic activity assays, bioinformatic sequence analysis","journal":"Biochemical and biophysical research communications","confidence":"Medium","confidence_rationale":"Tier 1 / Moderate — active-site mutagenesis with functional readout, single lab, multiple mutants tested","pmids":["16095562"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2006,"finding":"Peptidomimetic compounds based on a 1,4-benzodiazepine scaffold with a C-terminal aspartyl aldehyde provide significant inhibitory activity against FP2B (and FP2A), confirming FP2B is a druggable cysteine protease target.","method":"Enzyme inhibition assays with recombinant FP2B using synthetic peptidomimetic inhibitors","journal":"Journal of medicinal chemistry","confidence":"Low","confidence_rationale":"Tier 3 / Weak — inhibitor activity assay, single lab, mechanistic detail limited to inhibition potency","pmids":["16722625"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2013,"finding":"FP2B (falcipain 2/2') is a component of an ~200-kDa protein complex in the food vacuole that also contains plasmepsin II, plasmepsin IV, histo aspartic protease, and heme detoxification protein (HDP); this complex is required for hemoglobin degradation and hemozoin formation. Co-immunoprecipitation followed by mass spectrometry, gel filtration co-elution, glycerol gradient cosedimentation, and in vitro protein interaction analyses confirmed the associations. An in vitro assay showed that falcipain 2/2' and HDP together efficiently convert hemoglobin to hemozoin.","method":"Co-immunoprecipitation with mass spectrometry, gel filtration, glycerol gradient sedimentation, in vitro protein interaction assay, in vitro Hb-to-Hz conversion assay","journal":"Proceedings of the National Academy of Sciences of the United States of America","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple orthogonal biochemical methods (Co-IP/MS, gel filtration, sedimentation, in vitro reconstitution), single lab; note the paper uses 'falcipain 2/2'' without fully distinguishing FP2A from FP2B","pmids":["23471987"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]}],"current_model":"FP2B (PF3D7_1115300) is an active papain-family cysteine protease expressed in P. falciparum trophozoites that localizes to the food vacuole, hydrolyzes hemoglobin (pH optima 5.5–7.0), cleaves host erythrocyte ankyrin but not protein 4.1 in vitro, participates as part of an ~200-kDa multi-protein complex (with plasmepsins, HAP, and heme detoxification protein) that drives hemoglobin degradation and hemozoin formation, and has catalytically critical residues I85, S149, and A151 identified by mutagenesis."},"narrative":{"mechanistic_narrative":"FP2B (PF3D7_1115300) is an active papain-family cysteine protease expressed as an enzyme during the erythrocytic trophozoite stage of P. falciparum [PMID:16595182, PMID:16095562]. Its refolded mature protease domain is an acidic hemoglobinase with pH optima of 5.5–7.0, a reducing-agent requirement, and hemoglobin-hydrolyzing activity biochemically similar to the closely related isozyme falcipain-2 (97.5% sequence identity), from which it is distinguished by greater substitution tolerance at position 149 [PMID:16337629, PMID:16095562]. Active-site mutagenesis identified residues I85, S149, and A151 as structurally critical for catalysis, since double mutants at these positions abolished activity [PMID:16095562]. Beyond hemoglobin, recombinant FP2B selectively cleaves host erythrocyte ankyrin but not protein 4.1 in an inside-out-vesicle assay, implicating it in erythrocyte cytoskeletal proteolysis relevant to parasite egress [PMID:16595182]. FP2B is a component of an ~200-kDa food-vacuole complex that also contains plasmepsin II, plasmepsin IV, histo-aspartic protease, and heme detoxification protein (HDP), which together drive hemoglobin degradation and hemozoin formation; in vitro, FP2B and HDP efficiently convert hemoglobin to hemozoin [PMID:23471987]. FP2B is inhibited by 1,4-benzodiazepine-based peptidomimetic aspartyl-aldehyde compounds, establishing it as a druggable cysteine protease [PMID:16722625].","teleology":[{"year":2005,"claim":"Establishing that the FP2B protease domain is an autonomous, active hemoglobinase answered whether this near-identical paralog of falcipain-2 retains catalytic hemoglobin-degrading function.","evidence":"Recombinant mature domain expressed and refolded in E. coli, assayed against peptide substrates and hemoglobin with inhibitor profiling","pmids":["16337629"],"confidence":"Medium","gaps":["Activity shown only in vitro on isolated recombinant domain, not in the native food-vacuole context","Contribution of FP2B versus falcipain-2 to in-cell hemoglobin turnover not resolved"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2005,"claim":"Active-site mutagenesis defined which residues are catalytically essential and revealed a functional divergence from the nearly identical isozyme FP2A despite 97.5% identity.","evidence":"Site-directed mutagenesis (S149A/I85M and S149A/A151D double mutants) with enzymatic activity readout and sequence analysis","pmids":["16095562"],"confidence":"Medium","gaps":["Single-residue contributions not fully separated within double mutants","No structural model to explain differential position-149 tolerance versus FP2A"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2006,"claim":"Orthogonal enzymatic assays confirmed the full recombinant enzyme is proteolytically active and stage-restricted, anchoring FP2B as a trophozoite-expressed cysteine protease.","evidence":"Recombinant bacterial expression, fluorescent peptide cleavage, in-gel gelatin zymography, and RT-PCR staging","pmids":["16595182"],"confidence":"Medium","gaps":["No demonstration of native protein localization or endogenous activity","Physiological substrate spectrum not defined by these assays alone"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2006,"claim":"A selective erythrocyte-substrate cleavage assay tested whether FP2B acts on host cytoskeletal proteins, linking it to a potential egress-related role.","evidence":"Erythrocyte inside-out-vesicle assay with recombinant FP2B showing cleavage of ankyrin but not protein 4.1","pmids":["16595182"],"confidence":"Medium","gaps":["Single in vitro method without genetic confirmation of an egress role","Cleavage of ankyrin in intact infected cells not demonstrated"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2006,"claim":"Inhibitor profiling established that FP2B is chemically tractable, validating it as an antimalarial target.","evidence":"Enzyme inhibition assays of recombinant FP2B with 1,4-benzodiazepine aspartyl-aldehyde peptidomimetics","pmids":["16722625"],"confidence":"Low","gaps":["Mechanistic detail limited to inhibition potency, no structural basis of inhibition","Selectivity over FP2A not resolved","No cellular or parasite-killing correlation reported"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2013,"claim":"Complex characterization placed FP2B within a food-vacuole hemoglobin-degradation machine, connecting its protease activity to hemozoin biogenesis.","evidence":"Co-IP/MS, gel filtration co-elution, glycerol gradient sedimentation, and in vitro Hb-to-Hz conversion of the ~200-kDa complex containing plasmepsin II/IV, HAP, and HDP","pmids":["23471987"],"confidence":"Medium","gaps":["The study uses 'falcipain 2/2'' and does not fully distinguish FP2A from FP2B within the complex","Stoichiometry and direct FP2B contacts within the complex not defined"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":null,"claim":"The endogenous, gene-specific role of FP2B distinct from its near-identical paralog FP2A remains unresolved.","evidence":"","pmids":[],"confidence":"Low","gaps":["No genetic knockout/knockdown phenotype specific to FP2B in the corpus","In-cell localization and native function separated from FP2A not established","No structure of FP2B"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]}],"mechanism_profile":{"molecular_activity":[],"localization":[],"pathway":[],"complexes":["~200-kDa food-vacuole hemoglobin-degradation complex (with plasmepsin II, plasmepsin IV, histo-aspartic protease, and heme detoxification protein) [#5]"],"partners":["PLASMEPSIN II [#5]","PLASMEPSIN IV [#5]","HISTO-ASPARTIC PROTEASE (HAP) [#5]","HEME DETOXIFICATION PROTEIN (HDP) [#5]"],"other_free_text":["Catalytically critical residues I85, S149, and A151 identified by mutagenesis [#3]","pH optima 5.5–7.0 with reducing-agent requirement [#2]","Cleaves host erythrocyte ankyrin but not protein 4.1 in vitro [#1]","Inhibited by 1,4-benzodiazepine aspartyl-aldehyde peptidomimetics [#4]","97.5% sequence identity to the isozyme FP2A but more tolerant to substitution at position 149 [#3]","The 2013 complex study labels the component 'falcipain 2/2'' and does not fully separate FP2B from FP2A [#5]"]}},"prefetch_data":{},"corpus_meta":[{"pmid":"15070727","id":"PMC_15070727","title":"Gene disruption confirms a critical role for the cysteine protease falcipain-2 in hemoglobin hydrolysis by Plasmodium falciparum.","date":"2004","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/15070727","citation_count":245,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["curated_ref"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"23471987","id":"PMC_23471987","title":"Protein complex directs hemoglobin-to-hemozoin formation in Plasmodium falciparum.","date":"2013","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/23471987","citation_count":131,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["curated_ref"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"16890302","id":"PMC_16890302","title":"Gene disruptions demonstrate independent roles for the four falcipain cysteine proteases of Plasmodium falciparum.","date":"2006","source":"Molecular and biochemical parasitology","url":"https://pubmed.ncbi.nlm.nih.gov/16890302","citation_count":125,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["curated_ref"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"16722625","id":"PMC_16722625","title":"Novel peptidomimetic cysteine protease inhibitors as potential antimalarial agents.","date":"2006","source":"Journal of medicinal chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/16722625","citation_count":57,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["FP2B"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"16337629","id":"PMC_16337629","title":"Plasmodium falciparum: biochemical characterization of the cysteine protease falcipain-2'.","date":"2005","source":"Experimental parasitology","url":"https://pubmed.ncbi.nlm.nih.gov/16337629","citation_count":53,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["curated_ref"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"16095562","id":"PMC_16095562","title":"Characterization of amino acid variation at strategic positions in parasite and human proteases for selective inhibition of falcipains in Plasmodium falciparum.","date":"2005","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/16095562","citation_count":10,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["FP2B"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"16595182","id":"PMC_16595182","title":"Cloning and characterization of Plasmodium falciparum cysteine protease, falcipain-2B.","date":"2006","source":"Blood cells, molecules & diseases","url":"https://pubmed.ncbi.nlm.nih.gov/16595182","citation_count":9,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["curated_ref"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":5886,"output_tokens":1789,"usd":0.022246,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":6888,"output_tokens":2970,"usd":0.054345,"stage2_stop_reason":"end_turn"},"total_usd":0.076591,"stage1_batch_id":"msgbatch_01DisAaTypwmC8QwdMBuePHo","stage2_batch_id":"msgbatch_0184zxZTqxiEfdYYfPMfkqjb","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2006,\n      \"finding\": \"Recombinant FP2B expressed in bacteria exhibits cysteine protease activity, demonstrated by cleavage of a fluorescent peptide substrate and in-gel gelatin zymography. FP2B is transcribed at the trophozoite stage of P. falciparum.\",\n      \"method\": \"Recombinant protein expression in bacteria, fluorescent peptide substrate cleavage assay, in-gel gelatin zymography, RT-PCR\",\n      \"journal\": \"Blood cells, molecules & diseases\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct enzymatic assay with two orthogonal methods (fluorescent substrate + zymography), single lab\",\n      \"pmids\": [\"16595182\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"Recombinant FP2B cleaves host erythrocyte ankyrin but not protein 4.1, as assessed by an erythrocyte inside-out-vesicle assay in vitro, suggesting a selective proteolytic role in parasite egress from red blood cells.\",\n      \"method\": \"Erythrocyte inside-out-vesicle assay with recombinant FP2B\",\n      \"journal\": \"Blood cells, molecules & diseases\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Weak — direct in vitro substrate cleavage assay, single lab, single method\",\n      \"pmids\": [\"16595182\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"The mature protease domain of FP2B (falcipain-2'), expressed in E. coli and refolded, is an active hemoglobinase with biochemical properties similar to falcipain-2: pH optima of 5.5–7.0, reducing agent requirement, and similar substrate preference. Both enzymes readily hydrolyze hemoglobin in vitro.\",\n      \"method\": \"Recombinant protein expression and refolding in E. coli, enzymatic activity assays with peptide substrates and hemoglobin, inhibitor profiling\",\n      \"journal\": \"Experimental parasitology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro reconstitution of enzymatic activity with multiple substrate assays, single lab\",\n      \"pmids\": [\"16337629\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"FP2B (PF11_0161) is highly expressed as an active enzyme during the erythrocytic stage. Site-directed mutagenesis of active-site-proximate residues (S149A/I85M and S149A/A151D double mutants) rendered FP2B completely inactive, identifying I85, S149, and A151 as structurally critical residues. FP2B is more tolerant to substitution at position 149 than FP2A despite 97.5% sequence identity between the two isozymes.\",\n      \"method\": \"Active-site mutagenesis, enzymatic activity assays, bioinformatic sequence analysis\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — active-site mutagenesis with functional readout, single lab, multiple mutants tested\",\n      \"pmids\": [\"16095562\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"Peptidomimetic compounds based on a 1,4-benzodiazepine scaffold with a C-terminal aspartyl aldehyde provide significant inhibitory activity against FP2B (and FP2A), confirming FP2B is a druggable cysteine protease target.\",\n      \"method\": \"Enzyme inhibition assays with recombinant FP2B using synthetic peptidomimetic inhibitors\",\n      \"journal\": \"Journal of medicinal chemistry\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — inhibitor activity assay, single lab, mechanistic detail limited to inhibition potency\",\n      \"pmids\": [\"16722625\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2013,\n      \"finding\": \"FP2B (falcipain 2/2') is a component of an ~200-kDa protein complex in the food vacuole that also contains plasmepsin II, plasmepsin IV, histo aspartic protease, and heme detoxification protein (HDP); this complex is required for hemoglobin degradation and hemozoin formation. Co-immunoprecipitation followed by mass spectrometry, gel filtration co-elution, glycerol gradient cosedimentation, and in vitro protein interaction analyses confirmed the associations. An in vitro assay showed that falcipain 2/2' and HDP together efficiently convert hemoglobin to hemozoin.\",\n      \"method\": \"Co-immunoprecipitation with mass spectrometry, gel filtration, glycerol gradient sedimentation, in vitro protein interaction assay, in vitro Hb-to-Hz conversion assay\",\n      \"journal\": \"Proceedings of the National Academy of Sciences of the United States of America\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple orthogonal biochemical methods (Co-IP/MS, gel filtration, sedimentation, in vitro reconstitution), single lab; note the paper uses 'falcipain 2/2'' without fully distinguishing FP2A from FP2B\",\n      \"pmids\": [\"23471987\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    }\n  ],\n  \"current_model\": \"FP2B (PF3D7_1115300) is an active papain-family cysteine protease expressed in P. falciparum trophozoites that localizes to the food vacuole, hydrolyzes hemoglobin (pH optima 5.5–7.0), cleaves host erythrocyte ankyrin but not protein 4.1 in vitro, participates as part of an ~200-kDa multi-protein complex (with plasmepsins, HAP, and heme detoxification protein) that drives hemoglobin degradation and hemozoin formation, and has catalytically critical residues I85, S149, and A151 identified by mutagenesis.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"FP2B (PF3D7_1115300) is an active papain-family cysteine protease expressed as an enzyme during the erythrocytic trophozoite stage of P. falciparum [#0, #3]. Its refolded mature protease domain is an acidic hemoglobinase with pH optima of 5.5\\u20137.0, a reducing-agent requirement, and hemoglobin-hydrolyzing activity biochemically similar to the closely related isozyme falcipain-2 (97.5% sequence identity), from which it is distinguished by greater substitution tolerance at position 149 [#2, #3]. Active-site mutagenesis identified residues I85, S149, and A151 as structurally critical for catalysis, since double mutants at these positions abolished activity [#3]. Beyond hemoglobin, recombinant FP2B selectively cleaves host erythrocyte ankyrin but not protein 4.1 in an inside-out-vesicle assay, implicating it in erythrocyte cytoskeletal proteolysis relevant to parasite egress [#1]. FP2B is a component of an ~200-kDa food-vacuole complex that also contains plasmepsin II, plasmepsin IV, histo-aspartic protease, and heme detoxification protein (HDP), which together drive hemoglobin degradation and hemozoin formation; in vitro, FP2B and HDP efficiently convert hemoglobin to hemozoin [#5]. FP2B is inhibited by 1,4-benzodiazepine-based peptidomimetic aspartyl-aldehyde compounds, establishing it as a druggable cysteine protease [#4].\",\n  \"teleology\": [\n    {\n      \"year\": 2005,\n      \"claim\": \"Establishing that the FP2B protease domain is an autonomous, active hemoglobinase answered whether this near-identical paralog of falcipain-2 retains catalytic hemoglobin-degrading function.\",\n      \"evidence\": \"Recombinant mature domain expressed and refolded in E. coli, assayed against peptide substrates and hemoglobin with inhibitor profiling\",\n      \"pmids\": [\"16337629\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Activity shown only in vitro on isolated recombinant domain, not in the native food-vacuole context\", \"Contribution of FP2B versus falcipain-2 to in-cell hemoglobin turnover not resolved\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Active-site mutagenesis defined which residues are catalytically essential and revealed a functional divergence from the nearly identical isozyme FP2A despite 97.5% identity.\",\n      \"evidence\": \"Site-directed mutagenesis (S149A/I85M and S149A/A151D double mutants) with enzymatic activity readout and sequence analysis\",\n      \"pmids\": [\"16095562\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single-residue contributions not fully separated within double mutants\", \"No structural model to explain differential position-149 tolerance versus FP2A\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2006,\n      \"claim\": \"Orthogonal enzymatic assays confirmed the full recombinant enzyme is proteolytically active and stage-restricted, anchoring FP2B as a trophozoite-expressed cysteine protease.\",\n      \"evidence\": \"Recombinant bacterial expression, fluorescent peptide cleavage, in-gel gelatin zymography, and RT-PCR staging\",\n      \"pmids\": [\"16595182\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No demonstration of native protein localization or endogenous activity\", \"Physiological substrate spectrum not defined by these assays alone\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2006,\n      \"claim\": \"A selective erythrocyte-substrate cleavage assay tested whether FP2B acts on host cytoskeletal proteins, linking it to a potential egress-related role.\",\n      \"evidence\": \"Erythrocyte inside-out-vesicle assay with recombinant FP2B showing cleavage of ankyrin but not protein 4.1\",\n      \"pmids\": [\"16595182\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single in vitro method without genetic confirmation of an egress role\", \"Cleavage of ankyrin in intact infected cells not demonstrated\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2006,\n      \"claim\": \"Inhibitor profiling established that FP2B is chemically tractable, validating it as an antimalarial target.\",\n      \"evidence\": \"Enzyme inhibition assays of recombinant FP2B with 1,4-benzodiazepine aspartyl-aldehyde peptidomimetics\",\n      \"pmids\": [\"16722625\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Mechanistic detail limited to inhibition potency, no structural basis of inhibition\", \"Selectivity over FP2A not resolved\", \"No cellular or parasite-killing correlation reported\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Complex characterization placed FP2B within a food-vacuole hemoglobin-degradation machine, connecting its protease activity to hemozoin biogenesis.\",\n      \"evidence\": \"Co-IP/MS, gel filtration co-elution, glycerol gradient sedimentation, and in vitro Hb-to-Hz conversion of the ~200-kDa complex containing plasmepsin II/IV, HAP, and HDP\",\n      \"pmids\": [\"23471987\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"The study uses 'falcipain 2/2'' and does not fully distinguish FP2A from FP2B within the complex\", \"Stoichiometry and direct FP2B contacts within the complex not defined\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"The endogenous, gene-specific role of FP2B distinct from its near-identical paralog FP2A remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No genetic knockout/knockdown phenotype specific to FP2B in the corpus\", \"In-cell localization and native function separated from FP2A not established\", \"No structure of FP2B\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0004197\", \"supporting_discovery_ids\": [0, 2, 3]},\n      {\"term_id\": \"GO:0004175\", \"supporting_discovery_ids\": [2, 5]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0020020\", \"supporting_discovery_ids\": [5]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"GO:0044245\", \"supporting_discovery_ids\": [2, 5]}\n    ],\n    \"complexes\": [\n      \"~200-kDa food-vacuole hemoglobin-degradation complex (with plasmepsin II, plasmepsin IV, histo-aspartic protease, and heme detoxification protein) [#5]\"\n    ],\n    \"partners\": [\n      \"plasmepsin II [#5]\",\n      \"plasmepsin IV [#5]\",\n      \"histo-aspartic protease (HAP) [#5]\",\n      \"heme detoxification protein (HDP) [#5]\"\n    ],\n    \"other_free_text\": [\n      \"Catalytically critical residues I85, S149, and A151 identified by mutagenesis [#3]\",\n      \"pH optima 5.5\\u20137.0 with reducing-agent requirement [#2]\",\n      \"Cleaves host erythrocyte ankyrin but not protein 4.1 in vitro [#1]\",\n      \"Inhibited by 1,4-benzodiazepine aspartyl-aldehyde peptidomimetics [#4]\",\n      \"97.5% sequence identity to the isozyme FP2A but more tolerant to substitution at position 149 [#3]\",\n      \"The 2013 complex study labels the component 'falcipain 2/2'' and does not fully separate FP2B from FP2A [#5]\"\n    ]\n  }\n}","audit_flag":null,"strains":[{"strain":"Pf3D7","gene_id":"PF3D7_1115300","name":null,"product":"cysteine proteinase falcipain 2b","uniprot_acc":null,"chromosome":null,"start_pos":null}],"gene_symbol":"FP2B","evaluation":null}