{"gene":"TG_289690","gene_symbol":"GAPDH1","run_date":"2026-09-08T14:08:30","timeline":{"discoveries":[{"year":2016,"finding":"Crystal structure of T. gondii GAPDH1 holoenzyme resolved at 2.25 Å resolution as a quaternary complex, providing molecular basis for structure-function analysis.","method":"X-ray crystallography (2.25 Å resolution crystal structure)","journal":"Molecular microbiology","confidence":"High","confidence_rationale":"Tier 1 / Strong — atomic-resolution crystal structure combined with functional mutagenesis and knockdown experiments in a single rigorous study","pmids":["27859784"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2016,"finding":"Knockdown of GAPDH1 expression and catalytic site disruption demonstrate that GAPDH1 is essential for intracellular replication, but glycolysis per se is not strictly essential.","method":"Inducible knockdown and catalytic site mutagenesis with intracellular replication phenotype readout","journal":"Molecular microbiology","confidence":"High","confidence_rationale":"Tier 2 / Moderate — two orthogonal loss-of-function approaches (knockdown + active-site disruption) with defined cellular phenotype in a single rigorous study","pmids":["27859784"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2016,"finding":"S-loop phosphorylation of GAPDH1 is a novel post-translational modification that critically regulates enzymatic activity, consistent with the S-loop being required for cofactor binding, allosteric activation, and oligomerization.","method":"Phosphorylation site identification combined with structural analysis and enzymatic activity assays; S-loop mutagenesis","journal":"Molecular microbiology","confidence":"High","confidence_rationale":"Tier 1 / Moderate — structural validation plus enzymatic activity assays and mutagenesis in a single study with multiple orthogonal methods","pmids":["27859784"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2016,"finding":"Upon egress, GAPDH1 translocates to the cortical membrane skeleton, and this translocation is mediated by the N-terminus, likely through palmitoylation, and is functionally decoupled from enzymatic activity and phosphorylation state.","method":"Live imaging of GAPDH1 localization during egress; N-terminal deletion/mutagenesis; palmitoylation inhibition","journal":"Molecular microbiology","confidence":"High","confidence_rationale":"Tier 2 / Moderate — direct localization experiment with functional consequence, N-terminal domain mutagenesis implicating palmitoylation, decoupling shown by phosphorylation/activity variants, single rigorous study","pmids":["27859784"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""}]}],"current_model":"T. gondii GAPDH1 is an essential multifunctional enzyme whose 2.25 Å crystal structure reveals a quaternary holoenzyme architecture; S-loop phosphorylation allosterically regulates its glycolytic activity, cofactor binding, and oligomerization, while a separate N-terminal palmitoylation-dependent mechanism mediates its egress-triggered translocation to the cortical membrane skeleton — with glycolysis and cortical association being functionally decoupled post-translational events."},"narrative":{"mechanistic_narrative":"Toxoplasma gondii GAPDH1 is an essential multifunctional glycolytic enzyme required for intracellular replication, though its glycolytic catalytic activity per se is not strictly essential [PMID:27859784]. A 2.25 Å crystal structure resolves GAPDH1 as a quaternary holoenzyme, providing the framework for its structure-function analysis [PMID:27859784]. Enzymatic activity is regulated by S-loop phosphorylation, a post-translational modification acting through an S-loop required for cofactor binding, allosteric activation, and oligomerization [PMID:27859784]. Independently of its catalytic and phosphorylation state, GAPDH1 translocates to the cortical membrane skeleton upon egress via an N-terminal, palmitoylation-dependent mechanism, establishing that its glycolytic and cortical-association functions are decoupled post-translational events [PMID:27859784]. Beyond these findings from a single structural and functional study, no further mechanistic detail has been characterized in the available corpus.","teleology":[{"year":2016,"claim":"Resolving whether GAPDH1's essentiality derives from glycolysis established that the enzyme is required for intracellular replication independent of its strict glycolytic role.","evidence":"inducible knockdown plus catalytic-site mutagenesis with intracellular replication readout in T. gondii","pmids":["27859784"],"confidence":"High","gaps":["the non-glycolytic function accounting for essentiality is not defined","no identification of downstream metabolic or structural pathways affected by knockdown"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2016,"claim":"The holoenzyme crystal structure provided the atomic framework needed to interpret how modifications and mutations alter GAPDH1 function.","evidence":"X-ray crystallography at 2.25 Å resolution of the T. gondii GAPDH1 quaternary complex","pmids":["27859784"],"confidence":"High","gaps":["no structural model of the phosphorylated versus unphosphorylated S-loop state","no structure of the palmitoylated N-terminus or membrane-associated form"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2016,"claim":"Identifying S-loop phosphorylation defined a post-translational switch coupling cofactor binding, allosteric activation, and oligomerization to enzymatic output.","evidence":"phosphosite identification with structural analysis, enzymatic activity assays, and S-loop mutagenesis in T. gondii","pmids":["27859784"],"confidence":"High","gaps":["the kinase responsible for S-loop phosphorylation is not identified","physiological conditions triggering phosphorylation are undefined"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2016,"claim":"Live imaging of egress-triggered relocalization established a second, catalytically independent function: N-terminal palmitoylation-dependent recruitment to the cortical membrane skeleton.","evidence":"live imaging of GAPDH1 during egress, N-terminal deletion/mutagenesis, and palmitoylation inhibition in T. gondii","pmids":["27859784"],"confidence":"High","gaps":["the palmitoyltransferase and modified residue are not identified","the functional consequence of cortical association during egress is not defined","the signal linking egress to translocation is unknown"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":null,"claim":"The non-glycolytic function underlying GAPDH1 essentiality and the role of its egress-triggered cortical association remain unresolved.","evidence":"","pmids":[],"confidence":"High","gaps":["no mechanistic link between cortical localization and replication essentiality","no identification of the regulatory kinase or palmitoyltransferase","no characterization across T. gondii strains"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]}],"mechanism_profile":{"molecular_activity":[],"localization":[],"pathway":[],"complexes":["GAPDH1 quaternary holoenzyme"],"partners":[],"other_free_text":["Essential for intracellular replication independent of strict glycolytic activity [#1]","S-loop phosphorylation regulates cofactor binding, allosteric activation, and oligomerization [#2]","N-terminal palmitoylation-dependent translocation to the cortical membrane skeleton upon egress, decoupled from enzymatic activity and phosphorylation state [#3]"]},"strain_conflicts":[]},"prefetch_data":{},"corpus_meta":[{"pmid":"27859784","id":"PMC_27859784","title":"Membrane skeletal association and post-translational allosteric regulation of Toxoplasma gondii GAPDH1.","date":"2016","source":"Molecular microbiology","url":"https://pubmed.ncbi.nlm.nih.gov/27859784","citation_count":15,"is_preprint":false},{"pmid":"41230256","id":"PMC_41230256","title":"Harnessing antigenic proteins of Toxoplasma gondii for efficient diagnosis: a study of promising candidates.","date":"2025","source":"Journal of parasitic diseases : official organ of the Indian Society for Parasitology","url":"https://pubmed.ncbi.nlm.nih.gov/41230256","citation_count":0,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":3306,"output_tokens":1005,"usd":0.012497,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":5941,"output_tokens":2071,"usd":0.04074,"stage2_stop_reason":"end_turn"},"total_usd":0.053237,"stage1_batch_id":"msgbatch_015xmQKDRqnTh9anE4m7LiD6","stage2_batch_id":"msgbatch_01JLpkFTVhoQBNgqHCKFrVVU","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2016,\n      \"finding\": \"Crystal structure of T. gondii GAPDH1 holoenzyme resolved at 2.25 Å resolution as a quaternary complex, providing molecular basis for structure-function analysis.\",\n      \"method\": \"X-ray crystallography (2.25 Å resolution crystal structure)\",\n      \"journal\": \"Molecular microbiology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — atomic-resolution crystal structure combined with functional mutagenesis and knockdown experiments in a single rigorous study\",\n      \"pmids\": [\"27859784\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"Knockdown of GAPDH1 expression and catalytic site disruption demonstrate that GAPDH1 is essential for intracellular replication, but glycolysis per se is not strictly essential.\",\n      \"method\": \"Inducible knockdown and catalytic site mutagenesis with intracellular replication phenotype readout\",\n      \"journal\": \"Molecular microbiology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — two orthogonal loss-of-function approaches (knockdown + active-site disruption) with defined cellular phenotype in a single rigorous study\",\n      \"pmids\": [\"27859784\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"S-loop phosphorylation of GAPDH1 is a novel post-translational modification that critically regulates enzymatic activity, consistent with the S-loop being required for cofactor binding, allosteric activation, and oligomerization.\",\n      \"method\": \"Phosphorylation site identification combined with structural analysis and enzymatic activity assays; S-loop mutagenesis\",\n      \"journal\": \"Molecular microbiology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — structural validation plus enzymatic activity assays and mutagenesis in a single study with multiple orthogonal methods\",\n      \"pmids\": [\"27859784\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2016,\n      \"finding\": \"Upon egress, GAPDH1 translocates to the cortical membrane skeleton, and this translocation is mediated by the N-terminus, likely through palmitoylation, and is functionally decoupled from enzymatic activity and phosphorylation state.\",\n      \"method\": \"Live imaging of GAPDH1 localization during egress; N-terminal deletion/mutagenesis; palmitoylation inhibition\",\n      \"journal\": \"Molecular microbiology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct localization experiment with functional consequence, N-terminal domain mutagenesis implicating palmitoylation, decoupling shown by phosphorylation/activity variants, single rigorous study\",\n      \"pmids\": [\"27859784\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    }\n  ],\n  \"current_model\": \"T. gondii GAPDH1 is an essential multifunctional enzyme whose 2.25 Å crystal structure reveals a quaternary holoenzyme architecture; S-loop phosphorylation allosterically regulates its glycolytic activity, cofactor binding, and oligomerization, while a separate N-terminal palmitoylation-dependent mechanism mediates its egress-triggered translocation to the cortical membrane skeleton — with glycolysis and cortical association being functionally decoupled post-translational events.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"Toxoplasma gondii GAPDH1 is an essential multifunctional glycolytic enzyme required for intracellular replication, though its glycolytic catalytic activity per se is not strictly essential [#1]. A 2.25 Å crystal structure resolves GAPDH1 as a quaternary holoenzyme, providing the framework for its structure-function analysis [#0]. Enzymatic activity is regulated by S-loop phosphorylation, a post-translational modification acting through an S-loop required for cofactor binding, allosteric activation, and oligomerization [#2]. Independently of its catalytic and phosphorylation state, GAPDH1 translocates to the cortical membrane skeleton upon egress via an N-terminal, palmitoylation-dependent mechanism, establishing that its glycolytic and cortical-association functions are decoupled post-translational events [#3]. Beyond these findings from a single structural and functional study, no further mechanistic detail has been characterized in the available corpus.\",\n  \"teleology\": [\n    {\n      \"year\": 2016,\n      \"claim\": \"Resolving whether GAPDH1's essentiality derives from glycolysis established that the enzyme is required for intracellular replication independent of its strict glycolytic role.\",\n      \"evidence\": \"inducible knockdown plus catalytic-site mutagenesis with intracellular replication readout in T. gondii\",\n      \"pmids\": [\"27859784\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"the non-glycolytic function accounting for essentiality is not defined\",\n        \"no identification of downstream metabolic or structural pathways affected by knockdown\"\n      ],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"The holoenzyme crystal structure provided the atomic framework needed to interpret how modifications and mutations alter GAPDH1 function.\",\n      \"evidence\": \"X-ray crystallography at 2.25 Å resolution of the T. gondii GAPDH1 quaternary complex\",\n      \"pmids\": [\"27859784\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"no structural model of the phosphorylated versus unphosphorylated S-loop state\",\n        \"no structure of the palmitoylated N-terminus or membrane-associated form\"\n      ],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Identifying S-loop phosphorylation defined a post-translational switch coupling cofactor binding, allosteric activation, and oligomerization to enzymatic output.\",\n      \"evidence\": \"phosphosite identification with structural analysis, enzymatic activity assays, and S-loop mutagenesis in T. gondii\",\n      \"pmids\": [\"27859784\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"the kinase responsible for S-loop phosphorylation is not identified\",\n        \"physiological conditions triggering phosphorylation are undefined\"\n      ],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2016,\n      \"claim\": \"Live imaging of egress-triggered relocalization established a second, catalytically independent function: N-terminal palmitoylation-dependent recruitment to the cortical membrane skeleton.\",\n      \"evidence\": \"live imaging of GAPDH1 during egress, N-terminal deletion/mutagenesis, and palmitoylation inhibition in T. gondii\",\n      \"pmids\": [\"27859784\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"the palmitoyltransferase and modified residue are not identified\",\n        \"the functional consequence of cortical association during egress is not defined\",\n        \"the signal linking egress to translocation is unknown\"\n      ],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"The non-glycolytic function underlying GAPDH1 essentiality and the role of its egress-triggered cortical association remain unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"High\",\n      \"gaps\": [\n        \"no mechanistic link between cortical localization and replication essentiality\",\n        \"no identification of the regulatory kinase or palmitoyltransferase\",\n        \"no characterization across T. gondii strains\"\n      ],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0004365\", \"supporting_discovery_ids\": [1, 2], \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]},\n      {\"term_id\": \"GO:0050661\", \"supporting_discovery_ids\": [2], \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005856\", \"supporting_discovery_ids\": [3], \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"GO:0006096\", \"supporting_discovery_ids\": [1, 2], \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]}\n    ],\n    \"complexes\": [\"GAPDH1 quaternary holoenzyme\"],\n    \"partners\": [],\n    \"other_free_text\": [\n      \"Essential for intracellular replication independent of strict glycolytic activity [#1]\",\n      \"S-loop phosphorylation regulates cofactor binding, allosteric activation, and oligomerization [#2]\",\n      \"N-terminal palmitoylation-dependent translocation to the cortical membrane skeleton upon egress, decoupled from enzymatic activity and phosphorylation state [#3]\"\n    ]\n  },\n  \"strain_conflicts\": []\n}","audit_flag":null,"strains":[{"strain":"ME49","gene_id":"TGME49_289690","name":"GAPDH1","product":"glyceraldehyde-3-phosphate dehydrogenase GAPDH1","uniprot_acc":"A0A125YJ40","chromosome":"TGME49_chrIX","start_pos":3380629},{"strain":"GT1","gene_id":"TGGT1_289690","name":null,"product":"glyceraldehyde-3-phosphate dehydrogenase GAPDH1","uniprot_acc":null,"chromosome":"TGGT1_chrIX","start_pos":3189155},{"strain":"RH88","gene_id":"TGRH88_014440","name":null,"product":"glyceraldehyde-3-phosphate dehydrogenase GAPDH1","uniprot_acc":null,"chromosome":"CM023090","start_pos":3444946},{"strain":"VEG","gene_id":"TGVEG_289690","name":null,"product":"glyceraldehyde-3-phosphate dehydrogenase GAPDH1","uniprot_acc":"A0A125YJ41","chromosome":"TGVEG_chrIX","start_pos":3415149}]}