{"gene":"TG_240640","gene_symbol":"","run_date":"2026-09-08T12:58:24","timeline":{"discoveries":[{"year":2005,"finding":"Two T. gondii CK1 isoforms (TgCK1alpha, 38 kDa; TgCK1beta, 49 kDa) were cloned and shown to be enzymatically active when immunoprecipitated from transiently transfected T. gondii parasites, and both were also expressed as recombinant proteins in E. coli with confirmed kinase activity.","method":"PCR cloning, transient transfection with FLAG-tagged constructs, immunoprecipitation-based kinase assay, recombinant expression in E. coli with biochemical characterization","journal":"Molecular and biochemical parasitology","confidence":"High","confidence_rationale":"Tier 1 / Moderate — in vitro enzymatic activity confirmed by both immunoprecipitation from parasites and recombinant E. coli expression, single lab but multiple orthogonal methods","pmids":["15811523"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2005,"finding":"TgCK1alpha is cytosolic, whereas TgCK1beta localizes predominantly to the plasma membrane; deletion mapping demonstrated that the C-terminal extension domain of TgCK1beta confers this membrane association.","method":"Subcellular localization of FLAG-tagged constructs in transiently transfected T. gondii, deletion mapping of the C-terminal domain","journal":"Molecular and biochemical parasitology","confidence":"High","confidence_rationale":"Tier 2 / Moderate — direct localization experiment combined with domain deletion mapping, single lab, two orthogonal methods","pmids":["15811523"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2005,"finding":"A native 38 kDa CK1 activity partially purified from T. gondii tachyzoites by ion-exchange and hydrophobic interaction chromatography closely resembles recombinant TgCK1alpha in biochemical and serological properties; no corresponding native TgCK1beta activity was detected in tachyzoite lysates.","method":"Partial purification from tachyzoite lysates by ion-exchange and hydrophobic interaction chromatography, kinase activity assay, serological characterization","journal":"Molecular and biochemical parasitology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct biochemical purification from native parasites with activity assay, single lab; TgCK1beta native activity result is explicitly negative","pmids":["15811523"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2005,"finding":"Purvalanol B and aminopurvalanol A selectively inhibit TgCK1alpha kinase activity, and the cell-permeable aminopurvalanol A also inhibits T. gondii parasite growth, indicating that structural differences between host and parasite CK1 can be pharmacologically exploited.","method":"Kinase inhibition assays with recombinant TgCK1alpha, parasite growth inhibition assays","journal":"Molecular and biochemical parasitology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vitro kinase inhibition assay plus cell-based growth inhibition, single lab, two complementary methods","pmids":["15811523"],"is_preprint":false,"attribution":[{"species":"Toxoplasma gondii","strain":""}]}],"current_model":"T. gondii encodes two CK1 isoforms: TgCK1alpha (38 kDa, cytosolic, the predominant native tachyzoite kinase) and TgCK1beta (49 kDa, plasma membrane-localized via its C-terminal extension), both enzymatically active; selective inhibition of TgCK1alpha by aminopurvalanol compounds also inhibits parasite growth, highlighting exploitable structural differences from host CK1."},"narrative":{"mechanistic_narrative":"Toxoplasma gondii encodes two enzymatically active casein kinase I isoforms, TgCK1alpha (38 kDa) and TgCK1beta (49 kDa), both confirmed to have kinase activity when immunoprecipitated from transfected parasites and when produced recombinantly in E. coli [PMID:15811523]. The two isoforms differ in localization: TgCK1alpha is cytosolic while TgCK1beta associates predominantly with the plasma membrane, an association conferred by its C-terminal extension domain [PMID:15811523]. TgCK1alpha appears to be the predominant native tachyzoite enzyme, as a native 38 kDa CK1 activity purified from tachyzoite lysates matches recombinant TgCK1alpha biochemically and serologically, with no corresponding native TgCK1beta activity detected [PMID:15811523]. TgCK1alpha activity is selectively inhibited by the purvalanol compounds purvalanol B and aminopurvalanol A, and cell-permeable aminopurvalanol A also blocks parasite growth, indicating structural differences from host CK1 that can be pharmacologically exploited [PMID:15811523]. Beyond this biochemical and pharmacological characterization, the physiological substrates and downstream pathways of the T. gondii CK1 isoforms have not been characterized in the available corpus.","teleology":[{"year":2005,"claim":"Establishing whether T. gondii encodes functional CK1 enzymes was the first step; cloning and biochemical characterization confirmed two active isoforms, defining CK1 as a genuine parasite kinase family.","evidence":"PCR cloning, FLAG-tagged transient transfection with immunoprecipitation kinase assay, and recombinant E. coli expression in T. gondii","pmids":["15811523"],"confidence":"High","gaps":["Physiological substrates of neither isoform identified","No genetic knockout to establish in vivo requirement"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2005,"claim":"To distinguish the two isoforms functionally, localization and domain mapping showed TgCK1alpha is cytosolic and TgCK1beta is plasma-membrane targeted via its C-terminal extension, indicating spatially segregated roles.","evidence":"Subcellular localization of FLAG-tagged constructs and C-terminal deletion mapping in transiently transfected T. gondii","pmids":["15811523"],"confidence":"High","gaps":["Mechanism of C-terminal membrane targeting (lipidation vs. protein interaction) unknown","Localization shown in transient transfection, not for endogenous protein"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2005,"claim":"Whether the recombinant isoforms reflect native parasite enzymes was addressed by purifying endogenous CK1 activity, revealing TgCK1alpha as the predominant native tachyzoite kinase with no detectable native TgCK1beta activity.","evidence":"Partial purification from tachyzoite lysates by ion-exchange and hydrophobic interaction chromatography with kinase and serological characterization","pmids":["15811523"],"confidence":"Medium","gaps":["Negative result for native TgCK1beta may reflect low abundance rather than absence","Stage-specific expression of TgCK1beta not tested"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":2005,"claim":"To test therapeutic exploitability, selective inhibition of TgCK1alpha by purvalanol compounds and growth inhibition by cell-permeable aminopurvalanol A established parasite CK1 as a druggable target distinct from host CK1.","evidence":"In vitro kinase inhibition assays with recombinant TgCK1alpha and parasite growth inhibition assays","pmids":["15811523"],"confidence":"Medium","gaps":["Growth inhibition not formally shown to act through CK1 in vivo","Selectivity over host CK1 inferred, not directly compared","Which isoform mediates the growth phenotype unresolved"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]},{"year":null,"claim":"The physiological substrates, downstream signaling pathways, and genetic requirement of each T. gondii CK1 isoform remain unknown.","evidence":"","pmids":[],"confidence":"Low","gaps":["No substrates identified for either isoform","No knockout/conditional depletion to establish essentiality","No structural model explaining host-parasite CK1 inhibitor selectivity"],"attribution":[{"species":"Toxoplasma gondii","strain":""}]}],"mechanism_profile":{"molecular_activity":[],"localization":[],"pathway":[],"complexes":[],"partners":[],"other_free_text":["TgCK1alpha is the predominant native tachyzoite CK1 activity; no native TgCK1beta activity detected in tachyzoite lysates [#2].","Plasma membrane association of TgCK1beta is conferred by its C-terminal extension domain [#1].","TgCK1alpha is selectively inhibited by purvalanol B and aminopurvalanol A; cell-permeable aminopurvalanol A inhibits parasite growth [#3]."]},"strain_conflicts":[]},"prefetch_data":{},"corpus_meta":[{"pmid":"15811523","id":"PMC_15811523","title":"Characterization of two T. gondii CK1 isoforms.","date":"2005","source":"Molecular and biochemical parasitology","url":"https://pubmed.ncbi.nlm.nih.gov/15811523","citation_count":26,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":3064,"output_tokens":1135,"usd":0.013109,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":6082,"output_tokens":2218,"usd":0.04293,"stage2_stop_reason":"end_turn"},"total_usd":0.056039,"stage1_batch_id":"msgbatch_01TEPNZ4MCzJ7xtsNwosiDRk","stage2_batch_id":"msgbatch_01FQShM7n1sfCr4pWTDShmki","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2005,\n      \"finding\": \"Two T. gondii CK1 isoforms (TgCK1alpha, 38 kDa; TgCK1beta, 49 kDa) were cloned and shown to be enzymatically active when immunoprecipitated from transiently transfected T. gondii parasites, and both were also expressed as recombinant proteins in E. coli with confirmed kinase activity.\",\n      \"method\": \"PCR cloning, transient transfection with FLAG-tagged constructs, immunoprecipitation-based kinase assay, recombinant expression in E. coli with biochemical characterization\",\n      \"journal\": \"Molecular and biochemical parasitology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — in vitro enzymatic activity confirmed by both immunoprecipitation from parasites and recombinant E. coli expression, single lab but multiple orthogonal methods\",\n      \"pmids\": [\"15811523\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"TgCK1alpha is cytosolic, whereas TgCK1beta localizes predominantly to the plasma membrane; deletion mapping demonstrated that the C-terminal extension domain of TgCK1beta confers this membrane association.\",\n      \"method\": \"Subcellular localization of FLAG-tagged constructs in transiently transfected T. gondii, deletion mapping of the C-terminal domain\",\n      \"journal\": \"Molecular and biochemical parasitology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct localization experiment combined with domain deletion mapping, single lab, two orthogonal methods\",\n      \"pmids\": [\"15811523\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"A native 38 kDa CK1 activity partially purified from T. gondii tachyzoites by ion-exchange and hydrophobic interaction chromatography closely resembles recombinant TgCK1alpha in biochemical and serological properties; no corresponding native TgCK1beta activity was detected in tachyzoite lysates.\",\n      \"method\": \"Partial purification from tachyzoite lysates by ion-exchange and hydrophobic interaction chromatography, kinase activity assay, serological characterization\",\n      \"journal\": \"Molecular and biochemical parasitology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct biochemical purification from native parasites with activity assay, single lab; TgCK1beta native activity result is explicitly negative\",\n      \"pmids\": [\"15811523\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2005,\n      \"finding\": \"Purvalanol B and aminopurvalanol A selectively inhibit TgCK1alpha kinase activity, and the cell-permeable aminopurvalanol A also inhibits T. gondii parasite growth, indicating that structural differences between host and parasite CK1 can be pharmacologically exploited.\",\n      \"method\": \"Kinase inhibition assays with recombinant TgCK1alpha, parasite growth inhibition assays\",\n      \"journal\": \"Molecular and biochemical parasitology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vitro kinase inhibition assay plus cell-based growth inhibition, single lab, two complementary methods\",\n      \"pmids\": [\"15811523\"],\n      \"is_preprint\": false,\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    }\n  ],\n  \"current_model\": \"T. gondii encodes two CK1 isoforms: TgCK1alpha (38 kDa, cytosolic, the predominant native tachyzoite kinase) and TgCK1beta (49 kDa, plasma membrane-localized via its C-terminal extension), both enzymatically active; selective inhibition of TgCK1alpha by aminopurvalanol compounds also inhibits parasite growth, highlighting exploitable structural differences from host CK1.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"Toxoplasma gondii encodes two enzymatically active casein kinase I isoforms, TgCK1alpha (38 kDa) and TgCK1beta (49 kDa), both confirmed to have kinase activity when immunoprecipitated from transfected parasites and when produced recombinantly in E. coli [#0]. The two isoforms differ in localization: TgCK1alpha is cytosolic while TgCK1beta associates predominantly with the plasma membrane, an association conferred by its C-terminal extension domain [#1]. TgCK1alpha appears to be the predominant native tachyzoite enzyme, as a native 38 kDa CK1 activity purified from tachyzoite lysates matches recombinant TgCK1alpha biochemically and serologically, with no corresponding native TgCK1beta activity detected [#2]. TgCK1alpha activity is selectively inhibited by the purvalanol compounds purvalanol B and aminopurvalanol A, and cell-permeable aminopurvalanol A also blocks parasite growth, indicating structural differences from host CK1 that can be pharmacologically exploited [#3]. Beyond this biochemical and pharmacological characterization, the physiological substrates and downstream pathways of the T. gondii CK1 isoforms have not been characterized in the available corpus.\",\n  \"teleology\": [\n    {\n      \"year\": 2005,\n      \"claim\": \"Establishing whether T. gondii encodes functional CK1 enzymes was the first step; cloning and biochemical characterization confirmed two active isoforms, defining CK1 as a genuine parasite kinase family.\",\n      \"evidence\": \"PCR cloning, FLAG-tagged transient transfection with immunoprecipitation kinase assay, and recombinant E. coli expression in T. gondii\",\n      \"pmids\": [\"15811523\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Physiological substrates of neither isoform identified\", \"No genetic knockout to establish in vivo requirement\"],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"To distinguish the two isoforms functionally, localization and domain mapping showed TgCK1alpha is cytosolic and TgCK1beta is plasma-membrane targeted via its C-terminal extension, indicating spatially segregated roles.\",\n      \"evidence\": \"Subcellular localization of FLAG-tagged constructs and C-terminal deletion mapping in transiently transfected T. gondii\",\n      \"pmids\": [\"15811523\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Mechanism of C-terminal membrane targeting (lipidation vs. protein interaction) unknown\", \"Localization shown in transient transfection, not for endogenous protein\"],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"Whether the recombinant isoforms reflect native parasite enzymes was addressed by purifying endogenous CK1 activity, revealing TgCK1alpha as the predominant native tachyzoite kinase with no detectable native TgCK1beta activity.\",\n      \"evidence\": \"Partial purification from tachyzoite lysates by ion-exchange and hydrophobic interaction chromatography with kinase and serological characterization\",\n      \"pmids\": [\"15811523\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Negative result for native TgCK1beta may reflect low abundance rather than absence\", \"Stage-specific expression of TgCK1beta not tested\"],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": 2005,\n      \"claim\": \"To test therapeutic exploitability, selective inhibition of TgCK1alpha by purvalanol compounds and growth inhibition by cell-permeable aminopurvalanol A established parasite CK1 as a druggable target distinct from host CK1.\",\n      \"evidence\": \"In vitro kinase inhibition assays with recombinant TgCK1alpha and parasite growth inhibition assays\",\n      \"pmids\": [\"15811523\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Growth inhibition not formally shown to act through CK1 in vivo\", \"Selectivity over host CK1 inferred, not directly compared\", \"Which isoform mediates the growth phenotype unresolved\"],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"The physiological substrates, downstream signaling pathways, and genetic requirement of each T. gondii CK1 isoform remain unknown.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No substrates identified for either isoform\", \"No knockout/conditional depletion to establish essentiality\", \"No structural model explaining host-parasite CK1 inhibitor selectivity\"],\n      \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0004674\", \"supporting_discovery_ids\": [0, 2, 3], \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [1, 2], \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]},\n      {\"term_id\": \"GO:0005886\", \"supporting_discovery_ids\": [1], \"attribution\": [{\"species\": \"Toxoplasma gondii\", \"strain\": \"\"}]}\n    ],\n    \"pathway\": [],\n    \"complexes\": [],\n    \"partners\": [],\n    \"other_free_text\": [\n      \"TgCK1alpha is the predominant native tachyzoite CK1 activity; no native TgCK1beta activity detected in tachyzoite lysates [#2].\",\n      \"Plasma membrane association of TgCK1beta is conferred by its C-terminal extension domain [#1].\",\n      \"TgCK1alpha is selectively inhibited by purvalanol B and aminopurvalanol A; cell-permeable aminopurvalanol A inhibits parasite growth [#3].\"\n    ]\n  },\n  \"strain_conflicts\": []\n}","audit_flag":null,"strains":[{"strain":"ME49","gene_id":"TGME49_240640","name":null,"product":"casein kinase I","uniprot_acc":"A0A125YXT2","chromosome":"TGME49_chrVI","start_pos":1348593},{"strain":"GT1","gene_id":"TGGT1_240640","name":null,"product":"casein kinase I","uniprot_acc":null,"chromosome":"TGGT1_chrVI","start_pos":1321734},{"strain":"RH88","gene_id":"TGRH88_030510","name":null,"product":"casein kinase I","uniprot_acc":null,"chromosome":"CM023087","start_pos":2359096},{"strain":"VEG","gene_id":"TGVEG_240640","name":null,"product":"casein kinase I","uniprot_acc":"B9PPN2","chromosome":"TGVEG_chrVI","start_pos":1366087}]}