{"id":"e6f24bd9-0572-531f-9ba5-200ed928c7b8","stable_key":"548ab9d6-3a9b-5bed-879c-17d03813b636:b2-txnrd-mutants-upstream-retained","predicate":"retains","statement":"Engineered rat TXNRD1 terminal mutants retained one FAD per subunit and rapidly formed a thiolate-flavin charge-transfer signal upon NADPH titration.","claim_class":"mechanistic","status":"source_derived_draft","evidence_grade":"ungraded","direction":"neutral","is_public":true,"mechanism_event_id":"87680fc5-baaa-5bac-b18d-bf1c32f319f0","mechanism_event_label":"A defective terminal center can leave upstream flavin reduction intact.","subject":{"id":"3e52ad46-1a6e-52c4-b5d6-fda4a32b8725","slug":"rat-txnrd1-sec498cys","display_name":"Rat TXNRD1 Sec498Cys protein","entity_type_key":"protein_state"},"object":{"id":"e2cd7179-f218-54e8-9ce9-7a836ae35fac","slug":"fad","display_name":"FAD","entity_type_key":"small_molecule"},"evidence_count":1,"mechanism_event":{"id":"87680fc5-baaa-5bac-b18d-bf1c32f319f0","stable_key":"548ab9d6-3a9b-5bed-879c-17d03813b636:b2-txnrd-mutants-upstream-retained-event","event_type":"biochemical_relationship","label":"A defective terminal center can leave upstream flavin reduction intact.","description":"Engineered rat TXNRD1 terminal mutants retained one FAD per subunit and rapidly formed a thiolate-flavin charge-transfer signal upon NADPH titration.","status":"provisional","compartment":null,"participants":[{"entity":{"id":"e2cd7179-f218-54e8-9ce9-7a836ae35fac","slug":"fad","display_name":"FAD","entity_type_key":"small_molecule"},"role":"retained cofactor","stoichiometry":null,"state_label":"","sequence_order":0,"notes":""},{"entity":{"id":"4aba2a5e-8d06-5304-bb01-c0402b225a94","slug":"nadph","display_name":"NADPH","entity_type_key":"small_molecule"},"role":"reducing titrant","stoichiometry":null,"state_label":"","sequence_order":1,"notes":""},{"entity":{"id":"44c93642-76cb-5254-9edb-14a3020a5ae6","slug":"rat-txnrd1","display_name":"Rat thioredoxin reductase 1 / Txnrd1","entity_type_key":"protein"},"role":"wild-type comparator","stoichiometry":null,"state_label":"","sequence_order":2,"notes":""},{"entity":{"id":"3e52ad46-1a6e-52c4-b5d6-fda4a32b8725","slug":"rat-txnrd1-sec498cys","display_name":"Rat TXNRD1 Sec498Cys protein","entity_type_key":"protein_state"},"role":"subject","stoichiometry":null,"state_label":"","sequence_order":3,"notes":""}]},"contexts":[{"dimension":"availability_state","value_text":"machinery_impairment","comparator":null,"unit":null,"notes":"Imported condition classification; unverified.","entity":null},{"dimension":"cross_nutrient","value_text":"Distinct B2-cofactor and selenium-site functions cannot substitute for one another.","comparator":null,"unit":null,"notes":"","entity":null},{"dimension":"evidence_location","value_text":"Abstract","comparator":null,"unit":null,"notes":"","entity":null},{"dimension":"experimental_model","value_text":"Rat TXNRD1 Sec498Cys, Sec498Ser and C-terminal truncation expressed in E. coli; FAD analysis, NADPH titration and thioredoxin assays.","comparator":null,"unit":null,"notes":"","entity":null},{"dimension":"exposure","value_text":"Purified-enzyme assay","comparator":null,"unit":null,"notes":"","entity":null},{"dimension":"limitations","value_text":"Includes Sec498Cys, Sec498Ser and terminal truncation; engineered proteins, not nutrient-depleted animals.","comparator":null,"unit":null,"notes":"","entity":null},{"dimension":"nutrient_topic","value_text":"Riboflavin research collection; topical membership is not evidence of a direct dietary effect.","comparator":null,"unit":null,"notes":"","entity":{"slug":"riboflavin","display_name":"Riboflavin (vitamin B2)","entity_type_key":"small_molecule"}},{"dimension":"organism","value_text":"Rattus norvegicus protein expressed in Escherichia coli","comparator":null,"unit":null,"notes":"","entity":null},{"dimension":"plain_language","value_text":"A defective terminal center can leave upstream flavin reduction intact.","comparator":null,"unit":null,"notes":"","entity":null},{"dimension":"primary_references","value_text":"[zhong2000] Essential role of selenium in the catalytic activities of mammalian thioredoxin reductase revealed by characterization of recombinant enzymes with selenocysteine mutations. (2000). https://pubmed.ncbi.nlm.nih.gov/10849437/ DOI: 10.1074/jbc.m000690200","comparator":null,"unit":null,"notes":"","entity":null},{"dimension":"tissue_or_cell_type","value_text":"Purified recombinant enzyme; no intact tissue","comparator":null,"unit":null,"notes":"","entity":null},{"dimension":"trigger_kind","value_text":"machinery_impairment","comparator":null,"unit":null,"notes":"Imported condition classification; unverified.","entity":null}],"evidence":[{"id":"797ace77-27d5-5810-bf4f-d0fd604fcb75","evidence_kind":"source_excerpt","locator":"Lines 1374-1386","start_line":1374,"end_line":1386,"excerpt":"### b2-txnrd-mutants-upstream-retained\nEngineered rat TXNRD1 terminal mutants retained one FAD per subunit and rapidly formed a thiolate-flavin charge-transfer signal upon NADPH titration.\nCondition category: machinery_impairment\nnutrient_topic: Riboflavin research collection; topical membership is not evidence of a direct dietary effect.\nplain_language: A defective terminal center can leave upstream flavin reduction intact.\norganism: Rattus norvegicus protein expressed in Escherichia coli\ntissue_or_cell_type: Purified recombinant enzyme; no intact tissue\nexperimental_model: Rat TXNRD1 Sec498Cys, Sec498Ser and C-terminal truncation expressed in E. coli; FAD analysis, NADPH titration and thioredoxin assays.\nlimitations: Includes Sec498Cys, Sec498Ser and terminal truncation; engineered proteins, not nutrient-depleted animals.\nexposure: Purified-enzyme assay\ncross_nutrient: Distinct B2-cofactor and selenium-site functions cannot substitute for one another.\nevidence_location: Abstract\n[zhong2000] Essential role of selenium in the catalytic activities of mammalian thioredoxin reductase revealed by characterization of recombinant enzymes with selenocysteine mutations. 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