{"gene":"PF3D7_0915400","run_date":"2026-09-08T19:44:17","timeline":{"discoveries":[{"year":2018,"finding":"Hypomorphic (loss-of-function) mutations in PFK9 suppress the metabolic dysregulation and growth defect caused by loss of HAD2 in P. falciparum. Metabolic profiling confirmed reduced PFK9 function in suppressed strains, establishing PFK9 as a site of genetic metabolic plasticity that modulates glycolytic flux and downstream MEP pathway intermediate levels.","method":"Forward genetic suppressor screen (iterative culture under drug pressure), whole-genome sequencing of suppressor mutants, metabolic profiling of glycolytic and MEP pathway intermediates","journal":"mBio","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic epistasis confirmed by metabolic profiling, single lab, two orthogonal methods (genetics + metabolomics)","pmids":["30425143"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2020,"finding":"Mutations in PFK9 are associated with in vitro resistance to the sulfonamide glycoside PS-3, with resistant parasites showing significantly reduced flux through upper glycolysis, reduced ATP levels, increased flux into the pentose phosphate pathway, and reduced MEP pathway intermediates, indicating PFK9 activity controls the balance between glycolytic and alternative carbon routes.","method":"In vitro drug resistance selection, whole-genome sequencing of resistant parasites, metabolic flux analysis, metabolite profiling","journal":"mBio","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic and metabolic phenotyping with multiple orthogonal readouts, single lab","pmids":["33293381"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2016,"finding":"Kinetic modelling of P. falciparum glycolysis identified phosphofructokinase as having a high flux control coefficient, indicating PFK9 substantially controls glycolytic flux rate in the parasite.","method":"Detailed kinetic mathematical modelling of glycolysis with sensitivity (flux control coefficient) analysis","journal":"The FEBS journal","confidence":"Low","confidence_rationale":"Tier 4 / Weak — computational modelling only, no direct in vitro assay of PFK9 activity performed","pmids":["26648082"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2014,"finding":"Recombinant PFK9 protein and parasite extracts exhibit ATP-dependent phosphofructokinase activity, but no PPi-dependent activity was detected, establishing that PFK9 is a catalytically active, ATP-dependent enzyme despite containing plant-like PPi-PFK domain features. In P. berghei, the orthologous phosphofructokinase shows atypical allosteric regulation: AMP causes only marginal activation (unlike most PFKs), phosphoenolpyruvate activates (rather than inhibits) the enzyme, and ADP does not activate.","method":"In vitro enzymatic assay of recombinant PFK9 and parasite extracts with ATP and PPi as substrates; allosteric regulation assays in P. berghei","journal":"Malaria research and treatment","confidence":"Medium","confidence_rationale":"Tier 1 / Moderate — in vitro enzymatic reconstitution with substrate specificity testing; allosteric data from rodent ortholog cited as established prior work, single lab per cited reference","pmids":["25580350"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""},{"species":"Plasmodium berghei","strain":""}]},{"year":2021,"finding":"PFK9 (PF3D7_0915400) was identified as a putative substrate of PfCDPK4 by phosphoproteomic analysis of wild-type vs. cdpk4 knockout late gametocytes, with subsequent in vitro LC-MS/MS assay using recombinant PfCDPK4 and a synthetic PFK9-derived peptide showing significant phosphorylation, suggesting PFK9 is phosphorylated by PfCDPK4 during male gametogenesis.","method":"Mass spectrometry-based phosphoproteomics of cdpk4 knockout vs. wild-type gametocytes; in vitro kinase assay with recombinant PfCDPK4 and synthetic peptides","journal":"mBio","confidence":"Low","confidence_rationale":"Tier 3 / Weak — in vitro peptide phosphorylation assay is supportive but does not demonstrate phosphorylation of the full-length protein in situ or establish functional consequence for PFK9","pmids":["34724830"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2021,"finding":"PFK9 (PF3D7_0915400) was detected as a co-immunoprecipitation interactor of the K13 protein in P. falciparum (identified in the Gnädig et al. 2020 dataset and confirmed as the single protein overlapping with the BioID interactome of Birnbaum et al. 2020), suggesting a physical or proximal association between PFK9 and K13 in the parasite.","method":"Co-immunoprecipitation with anti-K13 monoclonal antibody; proximity labelling (BioID) by a separate group","journal":"FEMS microbiology reviews","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single Co-IP/BioID detection without functional validation of the PFK9–K13 interaction, reviewed rather than primary finding","pmids":["33095255"],"is_preprint":false,"attribution":[{"species":"Plasmodium falciparum","strain":""}]}],"current_model":"PFK9 (PF3D7_0915400) is a catalytically active, ATP-dependent phosphofructokinase that catalyzes the rate-limiting conversion of fructose-6-phosphate to fructose-1,6-bisphosphate in P. falciparum glycolysis; it has a high flux control coefficient over glycolytic output, and hypomorphic PFK9 mutations suppress the metabolic dysregulation caused by loss of HAD2, revealing PFK9 as a key node of metabolic plasticity linking glycolytic flux to the MEP isoprenoid pathway; additionally, PFK9 is a candidate substrate for phosphorylation by PfCDPK4 during male gametogenesis and has been detected in physical proximity to the artemisinin-resistance protein K13."},"narrative":{"mechanistic_narrative":"PFK9 (PF3D7_0915400) is the ATP-dependent 6-phosphofructokinase of Plasmodium falciparum and a principal control point of parasite glycolysis, catalyzing phosphorylation of fructose-6-phosphate to fructose-1,6-bisphosphate [PMID:25580350]. Despite carrying plant-like PPi-PFK domain features, recombinant PFK9 and parasite extracts display ATP-dependent activity with no detectable PPi-dependent activity, establishing it as a catalytically active, ATP-utilizing enzyme; the orthologous enzyme in P. berghei shows atypical allosteric behavior in which phosphoenolpyruvate activates rather than inhibits and AMP activates only marginally [PMID:25580350]. PFK9 exerts substantial control over glycolytic flux, and modulating its activity redistributes carbon between upper glycolysis, the pentose phosphate pathway, and the MEP isoprenoid pathway: hypomorphic PFK9 mutations suppress the metabolic dysregulation caused by loss of HAD2, while other PFK9 mutations confer resistance to the glycolytic inhibitor PS-3 with reduced upper-glycolytic flux, lower ATP, and diminished MEP intermediates [PMID:30425143, PMID:33293381]. This positions PFK9 as a node of metabolic plasticity linking glycolytic flux to downstream isoprenoid synthesis [PMID:30425143, PMID:33293381]. Beyond its glycolytic role, PFK9 has been identified as a candidate substrate for phosphorylation by PfCDPK4 during male gametogenesis and detected in physical proximity to the artemisinin-resistance protein K13, but neither association has been functionally characterized in the available corpus [PMID:34724830, PMID:33095255].","teleology":[{"year":2014,"claim":"Whether the Plasmodium enzyme with plant-like PPi-PFK domain features actually uses ATP or inorganic pyrophosphate was resolved by direct enzymatic reconstitution, defining PFK9 as a genuine ATP-dependent phosphofructokinase.","evidence":"In vitro enzymatic assay of recombinant PFK9 and parasite extracts with ATP vs. PPi substrates; allosteric regulation assays in the P. berghei ortholog","pmids":["25580350"],"confidence":"Medium","gaps":["No structural model of the P. falciparum enzyme reported","Allosteric data are from the P. berghei ortholog, not P. falciparum PFK9 directly","Physiological regulators controlling PFK9 activity in situ not defined"],"attribution":[{"species":"Plasmodium falciparum","strain":""},{"species":"Plasmodium berghei","strain":""}]},{"year":2016,"claim":"Whether phosphofructokinase is merely one glycolytic enzyme or a rate-controlling step was addressed by kinetic modelling, predicting a high flux control coefficient for the reaction PFK9 catalyzes.","evidence":"Detailed kinetic mathematical modelling of P. falciparum glycolysis with flux control coefficient sensitivity analysis","pmids":["26648082"],"confidence":"Low","gaps":["Computational prediction only, no direct in vitro or in vivo flux perturbation of PFK9","Does not identify which parameters set the control coefficient","Model not validated against a PFK9 knockdown or mutant"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2018,"claim":"Whether PFK9 activity is a tunable node connecting glycolysis to downstream metabolism was tested genetically, showing that reducing PFK9 function suppresses HAD2-loss dysregulation and remodels glycolytic and MEP intermediate levels.","evidence":"Forward genetic suppressor screen with whole-genome sequencing of suppressor mutants and metabolic profiling of glycolytic and MEP pathway intermediates","pmids":["30425143"],"confidence":"Medium","gaps":["Precise biochemical impact of the hypomorphic mutations on PFK9 kinetics not measured","Mechanism linking glycolytic flux to MEP intermediates not resolved at enzyme level","Single lab"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2020,"claim":"Whether PFK9 governs the partitioning of carbon among competing metabolic routes was addressed by drug-resistance selection, linking PFK9 mutations to reduced upper-glycolytic flux, lower ATP, increased pentose phosphate flux, and reduced MEP intermediates.","evidence":"In vitro PS-3 resistance selection with whole-genome sequencing, metabolic flux analysis, and metabolite profiling","pmids":["33293381"],"confidence":"Medium","gaps":["Whether resistance is due to altered PFK9 catalysis versus regulation not distinguished","Direct enzymatic characterization of resistant PFK9 alleles absent","Single lab"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2021,"claim":"Whether PFK9 is subject to signaling-driven post-translational control during sexual development was probed phosphoproteomically, identifying PFK9 as a candidate PfCDPK4 substrate in male gametogenesis.","evidence":"Phosphoproteomics of cdpk4 knockout vs. wild-type gametocytes plus in vitro kinase assay with recombinant PfCDPK4 and a synthetic PFK9-derived peptide","pmids":["34724830"],"confidence":"Low","gaps":["In vitro peptide assay does not demonstrate phosphorylation of full-length PFK9 in situ","Functional consequence of phosphorylation for PFK9 activity or gametogenesis unknown","Phosphosite not mapped in the intact protein context"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":2021,"claim":"Whether PFK9 physically associates with the artemisinin-resistance determinant K13 was raised by interactome data, detecting PFK9 as a shared Co-IP and BioID interactor of K13.","evidence":"Co-immunoprecipitation with anti-K13 monoclonal antibody and cross-referenced BioID proximity labelling, reported in a review","pmids":["33095255"],"confidence":"Low","gaps":["Single Co-IP/BioID detection without reciprocal validation or functional testing","Direct versus proximal/indirect association not distinguished","Biological role of any PFK9–K13 association undefined"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]},{"year":null,"claim":"How PFK9 activity is dynamically regulated in vivo — the functional consequences of PfCDPK4 phosphorylation, the significance of the K13 association, and the structural basis of its atypical allosteric behavior — remains unresolved.","evidence":"","pmids":[],"confidence":"Low","gaps":["No structural model or full-length in situ phosphorylation data","PFK9–K13 interaction lacks functional validation","Enzymatic characterization of disease/resistance-associated PFK9 alleles missing"],"attribution":[{"species":"Plasmodium falciparum","strain":""}]}],"mechanism_profile":{"molecular_activity":[],"localization":[],"pathway":[],"complexes":[],"partners":["PFCDPK4 (CANDIDATE KINASE, PHOSPHORYLATES PFK9-DERIVED PEPTIDE IN VITRO)","K13 (DETECTED AS CO-IP/BIOID PROXIMAL INTERACTOR)"],"other_free_text":["Catalytically active ATP-dependent 6-phosphofructokinase; no PPi-dependent activity detected despite plant-like PPi-PFK domain features [#3]","High flux control coefficient over parasite glycolysis predicted by kinetic modelling [#2]","Hypomorphic PFK9 mutations suppress HAD2-loss metabolic dysregulation [#0]","PFK9 mutations confer in vitro resistance to the glycolytic inhibitor PS-3 with reduced upper-glycolytic flux and shifted carbon partitioning [#1]","P. berghei ortholog shows atypical allosteric regulation: phosphoenolpyruvate activates, AMP only marginally activates, ADP does not activate [#3]"]}},"prefetch_data":{},"corpus_meta":[{"pmid":"26648082","id":"PMC_26648082","title":"Targeting glycolysis in the malaria parasite Plasmodium falciparum.","date":"2016","source":"The FEBS journal","url":"https://pubmed.ncbi.nlm.nih.gov/26648082","citation_count":36,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["ATP"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"30425143","id":"PMC_30425143","title":"Suppression of Drug Resistance Reveals a Genetic Mechanism of Metabolic Plasticity in Malaria Parasites.","date":"2018","source":"mBio","url":"https://pubmed.ncbi.nlm.nih.gov/30425143","citation_count":22,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["PFK9"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"33293381","id":"","title":"The Key Glycolytic Enzyme Phosphofructokinase Is Involved in Resistance to Antiplasmodial Glycosides.","date":"2020","source":"mBio","url":"https://pubmed.ncbi.nlm.nih.gov/33293381","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["PFK9"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"36103572","id":"","title":"GAPDH mediates drug resistance and metabolism in Plasmodium falciparum malaria parasites.","date":"2022","source":"PLoS pathogens","url":"https://pubmed.ncbi.nlm.nih.gov/36103572","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["PFK9"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"34724830","id":"","title":"Plasmodium falciparum Calcium-Dependent Protein Kinase 4 is Critical for Male Gametogenesis and Transmission to the Mosquito Vector.","date":"2021","source":"mBio","url":"https://pubmed.ncbi.nlm.nih.gov/34724830","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["PFK9"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"36154191","id":"","title":"Plasmodium falciparum CRK5 Is Critical for Male Gametogenesis and Infection of the Mosquito.","date":"2022","source":"mBio","url":"https://pubmed.ncbi.nlm.nih.gov/36154191","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["PFK9"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"31205418","id":"","title":"Haloacid Dehalogenase Proteins: Novel Mediators of Metabolic Plasticity in Plasmodium falciparum.","date":"2019","source":"Microbiology insights","url":"https://pubmed.ncbi.nlm.nih.gov/31205418","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["PFK9"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"25580350","id":"","title":"Exploiting unique structural and functional properties of malarial glycolytic enzymes for antimalarial drug development.","date":"2014","source":"Malaria research and treatment","url":"https://pubmed.ncbi.nlm.nih.gov/25580350","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["PFK9"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false},{"pmid":"33095255","id":"","title":"Artemisinin susceptibility in the malaria parasite Plasmodium falciparum: propellers, adaptor proteins and the need for cellular healing.","date":"2021","source":"FEMS microbiology reviews","url":"https://pubmed.ncbi.nlm.nih.gov/33095255","citation_count":0,"is_preprint":false,"source_species":["Pf"],"role":"primary","match_terms":["PFK9"],"competing_organism_flag":false,"competing_organism_match":"","competing_organism_only":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":11266,"output_tokens":1865,"usd":0.030887,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":7088,"output_tokens":3368,"usd":0.05982,"stage2_stop_reason":"end_turn"},"total_usd":0.090707,"stage1_batch_id":"msgbatch_01SRp5hyRc82zNjddxyCCXGk","stage2_batch_id":"msgbatch_01K3BTmBazRo3pRrMvyaPMTY","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2018,\n      \"finding\": \"Hypomorphic (loss-of-function) mutations in PFK9 suppress the metabolic dysregulation and growth defect caused by loss of HAD2 in P. falciparum. Metabolic profiling confirmed reduced PFK9 function in suppressed strains, establishing PFK9 as a site of genetic metabolic plasticity that modulates glycolytic flux and downstream MEP pathway intermediate levels.\",\n      \"method\": \"Forward genetic suppressor screen (iterative culture under drug pressure), whole-genome sequencing of suppressor mutants, metabolic profiling of glycolytic and MEP pathway intermediates\",\n      \"journal\": \"mBio\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic epistasis confirmed by metabolic profiling, single lab, two orthogonal methods (genetics + metabolomics)\",\n      \"pmids\": [\"30425143\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"Mutations in PFK9 are associated with in vitro resistance to the sulfonamide glycoside PS-3, with resistant parasites showing significantly reduced flux through upper glycolysis, reduced ATP levels, increased flux into the pentose phosphate pathway, and reduced MEP pathway intermediates, indicating PFK9 activity controls the balance between glycolytic and alternative carbon routes.\",\n      \"method\": \"In vitro drug resistance selection, whole-genome sequencing of resistant parasites, metabolic flux analysis, metabolite profiling\",\n      \"journal\": \"mBio\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic and metabolic phenotyping with multiple orthogonal readouts, single lab\",\n      \"pmids\": [\"33293381\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"Kinetic modelling of P. falciparum glycolysis identified phosphofructokinase as having a high flux control coefficient, indicating PFK9 substantially controls glycolytic flux rate in the parasite.\",\n      \"method\": \"Detailed kinetic mathematical modelling of glycolysis with sensitivity (flux control coefficient) analysis\",\n      \"journal\": \"The FEBS journal\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 4 / Weak — computational modelling only, no direct in vitro assay of PFK9 activity performed\",\n      \"pmids\": [\"26648082\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"Recombinant PFK9 protein and parasite extracts exhibit ATP-dependent phosphofructokinase activity, but no PPi-dependent activity was detected, establishing that PFK9 is a catalytically active, ATP-dependent enzyme despite containing plant-like PPi-PFK domain features. In P. berghei, the orthologous phosphofructokinase shows atypical allosteric regulation: AMP causes only marginal activation (unlike most PFKs), phosphoenolpyruvate activates (rather than inhibits) the enzyme, and ADP does not activate.\",\n      \"method\": \"In vitro enzymatic assay of recombinant PFK9 and parasite extracts with ATP and PPi as substrates; allosteric regulation assays in P. berghei\",\n      \"journal\": \"Malaria research and treatment\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro enzymatic reconstitution with substrate specificity testing; allosteric data from rodent ortholog cited as established prior work, single lab per cited reference\",\n      \"pmids\": [\"25580350\"],\n      \"is_preprint\": false,\n      \"attribution\": [\n        {\"species\": \"Plasmodium falciparum\", \"strain\": \"\"},\n        {\"species\": \"Plasmodium berghei\", \"strain\": \"\"}\n      ]\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"PFK9 (PF3D7_0915400) was identified as a putative substrate of PfCDPK4 by phosphoproteomic analysis of wild-type vs. cdpk4 knockout late gametocytes, with subsequent in vitro LC-MS/MS assay using recombinant PfCDPK4 and a synthetic PFK9-derived peptide showing significant phosphorylation, suggesting PFK9 is phosphorylated by PfCDPK4 during male gametogenesis.\",\n      \"method\": \"Mass spectrometry-based phosphoproteomics of cdpk4 knockout vs. wild-type gametocytes; in vitro kinase assay with recombinant PfCDPK4 and synthetic peptides\",\n      \"journal\": \"mBio\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — in vitro peptide phosphorylation assay is supportive but does not demonstrate phosphorylation of the full-length protein in situ or establish functional consequence for PFK9\",\n      \"pmids\": [\"34724830\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"PFK9 (PF3D7_0915400) was detected as a co-immunoprecipitation interactor of the K13 protein in P. falciparum (identified in the Gnädig et al. 2020 dataset and confirmed as the single protein overlapping with the BioID interactome of Birnbaum et al. 2020), suggesting a physical or proximal association between PFK9 and K13 in the parasite.\",\n      \"method\": \"Co-immunoprecipitation with anti-K13 monoclonal antibody; proximity labelling (BioID) by a separate group\",\n      \"journal\": \"FEMS microbiology reviews\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single Co-IP/BioID detection without functional validation of the PFK9–K13 interaction, reviewed rather than primary finding\",\n      \"pmids\": [\"33095255\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    }\n  ],\n  \"current_model\": \"PFK9 (PF3D7_0915400) is a catalytically active, ATP-dependent phosphofructokinase that catalyzes the rate-limiting conversion of fructose-6-phosphate to fructose-1,6-bisphosphate in P. falciparum glycolysis; it has a high flux control coefficient over glycolytic output, and hypomorphic PFK9 mutations suppress the metabolic dysregulation caused by loss of HAD2, revealing PFK9 as a key node of metabolic plasticity linking glycolytic flux to the MEP isoprenoid pathway; additionally, PFK9 is a candidate substrate for phosphorylation by PfCDPK4 during male gametogenesis and has been detected in physical proximity to the artemisinin-resistance protein K13.\"\n}\n```","stage2_raw":"```json\n{\n  \"mechanistic_narrative\": \"PFK9 (PF3D7_0915400) is the ATP-dependent 6-phosphofructokinase of Plasmodium falciparum and a principal control point of parasite glycolysis, catalyzing phosphorylation of fructose-6-phosphate to fructose-1,6-bisphosphate [#3]. Despite carrying plant-like PPi-PFK domain features, recombinant PFK9 and parasite extracts display ATP-dependent activity with no detectable PPi-dependent activity, establishing it as a catalytically active, ATP-utilizing enzyme; the orthologous enzyme in P. berghei shows atypical allosteric behavior in which phosphoenolpyruvate activates rather than inhibits and AMP activates only marginally [#3]. PFK9 exerts substantial control over glycolytic flux, and modulating its activity redistributes carbon between upper glycolysis, the pentose phosphate pathway, and the MEP isoprenoid pathway: hypomorphic PFK9 mutations suppress the metabolic dysregulation caused by loss of HAD2, while other PFK9 mutations confer resistance to the glycolytic inhibitor PS-3 with reduced upper-glycolytic flux, lower ATP, and diminished MEP intermediates [#0, #1]. This positions PFK9 as a node of metabolic plasticity linking glycolytic flux to downstream isoprenoid synthesis [#0, #1]. Beyond its glycolytic role, PFK9 has been identified as a candidate substrate for phosphorylation by PfCDPK4 during male gametogenesis and detected in physical proximity to the artemisinin-resistance protein K13, but neither association has been functionally characterized in the available corpus [#4, #5].\",\n  \"teleology\": [\n    {\n      \"year\": 2014,\n      \"claim\": \"Whether the Plasmodium enzyme with plant-like PPi-PFK domain features actually uses ATP or inorganic pyrophosphate was resolved by direct enzymatic reconstitution, defining PFK9 as a genuine ATP-dependent phosphofructokinase.\",\n      \"evidence\": \"In vitro enzymatic assay of recombinant PFK9 and parasite extracts with ATP vs. PPi substrates; allosteric regulation assays in the P. berghei ortholog\",\n      \"pmids\": [\"25580350\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No structural model of the P. falciparum enzyme reported\", \"Allosteric data are from the P. berghei ortholog, not P. falciparum PFK9 directly\", \"Physiological regulators controlling PFK9 activity in situ not defined\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}, {\"species\": \"Plasmodium berghei\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Whether phosphofructokinase is merely one glycolytic enzyme or a rate-controlling step was addressed by kinetic modelling, predicting a high flux control coefficient for the reaction PFK9 catalyzes.\",\n      \"evidence\": \"Detailed kinetic mathematical modelling of P. falciparum glycolysis with flux control coefficient sensitivity analysis\",\n      \"pmids\": [\"26648082\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Computational prediction only, no direct in vitro or in vivo flux perturbation of PFK9\", \"Does not identify which parameters set the control coefficient\", \"Model not validated against a PFK9 knockdown or mutant\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Whether PFK9 activity is a tunable node connecting glycolysis to downstream metabolism was tested genetically, showing that reducing PFK9 function suppresses HAD2-loss dysregulation and remodels glycolytic and MEP intermediate levels.\",\n      \"evidence\": \"Forward genetic suppressor screen with whole-genome sequencing of suppressor mutants and metabolic profiling of glycolytic and MEP pathway intermediates\",\n      \"pmids\": [\"30425143\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Precise biochemical impact of the hypomorphic mutations on PFK9 kinetics not measured\", \"Mechanism linking glycolytic flux to MEP intermediates not resolved at enzyme level\", \"Single lab\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Whether PFK9 governs the partitioning of carbon among competing metabolic routes was addressed by drug-resistance selection, linking PFK9 mutations to reduced upper-glycolytic flux, lower ATP, increased pentose phosphate flux, and reduced MEP intermediates.\",\n      \"evidence\": \"In vitro PS-3 resistance selection with whole-genome sequencing, metabolic flux analysis, and metabolite profiling\",\n      \"pmids\": [\"33293381\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether resistance is due to altered PFK9 catalysis versus regulation not distinguished\", \"Direct enzymatic characterization of resistant PFK9 alleles absent\", \"Single lab\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Whether PFK9 is subject to signaling-driven post-translational control during sexual development was probed phosphoproteomically, identifying PFK9 as a candidate PfCDPK4 substrate in male gametogenesis.\",\n      \"evidence\": \"Phosphoproteomics of cdpk4 knockout vs. wild-type gametocytes plus in vitro kinase assay with recombinant PfCDPK4 and a synthetic PFK9-derived peptide\",\n      \"pmids\": [\"34724830\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"In vitro peptide assay does not demonstrate phosphorylation of full-length PFK9 in situ\", \"Functional consequence of phosphorylation for PFK9 activity or gametogenesis unknown\", \"Phosphosite not mapped in the intact protein context\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Whether PFK9 physically associates with the artemisinin-resistance determinant K13 was raised by interactome data, detecting PFK9 as a shared Co-IP and BioID interactor of K13.\",\n      \"evidence\": \"Co-immunoprecipitation with anti-K13 monoclonal antibody and cross-referenced BioID proximity labelling, reported in a review\",\n      \"pmids\": [\"33095255\"],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"Single Co-IP/BioID detection without reciprocal validation or functional testing\", \"Direct versus proximal/indirect association not distinguished\", \"Biological role of any PFK9–K13 association undefined\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How PFK9 activity is dynamically regulated in vivo — the functional consequences of PfCDPK4 phosphorylation, the significance of the K13 association, and the structural basis of its atypical allosteric behavior — remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No structural model or full-length in situ phosphorylation data\", \"PFK9–K13 interaction lacks functional validation\", \"Enzymatic characterization of disease/resistance-associated PFK9 alleles missing\"],\n      \"attribution\": [{\"species\": \"Plasmodium falciparum\", \"strain\": \"\"}]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0003872\", \"supporting_discovery_ids\": [3]},\n      {\"term_id\": \"GO:0005524\", \"supporting_discovery_ids\": [3]},\n      {\"term_id\": \"GO:0016301\", \"supporting_discovery_ids\": [3]}\n    ],\n    \"localization\": [],\n    \"pathway\": [\n      {\"term_id\": \"GO:0006096\", \"supporting_discovery_ids\": [0, 1, 2, 3]},\n      {\"term_id\": \"GO:0006098\", \"supporting_discovery_ids\": [1]},\n      {\"term_id\": \"GO:0019288\", \"supporting_discovery_ids\": [0, 1]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\n      \"PfCDPK4 (candidate kinase, phosphorylates PFK9-derived peptide in vitro)\",\n      \"K13 (detected as Co-IP/BioID proximal interactor)\"\n    ],\n    \"other_free_text\": [\n      \"Catalytically active ATP-dependent 6-phosphofructokinase; no PPi-dependent activity detected despite plant-like PPi-PFK domain features [#3]\",\n      \"High flux control coefficient over parasite glycolysis predicted by kinetic modelling [#2]\",\n      \"Hypomorphic PFK9 mutations suppress HAD2-loss metabolic dysregulation [#0]\",\n      \"PFK9 mutations confer in vitro resistance to the glycolytic inhibitor PS-3 with reduced upper-glycolytic flux and shifted carbon partitioning [#1]\",\n      \"P. berghei ortholog shows atypical allosteric regulation: phosphoenolpyruvate activates, AMP only marginally activates, ADP does not activate [#3]\"\n    ]\n  }\n}\n```","audit_flag":null,"strains":[{"strain":"Pf3D7","gene_id":"PF3D7_0915400","name":null,"product":"ATP-dependent 6-phosphofructokinase","uniprot_acc":null,"chromosome":null,"start_pos":null},{"strain":"PbANKA","gene_id":"PBANKA_0816400","name":null,"product":"ATP-dependent 6-phosphofructokinase, putative","uniprot_acc":null,"chromosome":null,"start_pos":null},{"strain":"PkH","gene_id":"PKNH_0713400","name":null,"product":"ATP-dependent 6-phosphofructokinase, putative","uniprot_acc":null,"chromosome":null,"start_pos":null}],"gene_symbol":"PFK9","evaluation":null}