{"id":"2a72967d-3090-5fde-91f4-faa278a32e06","stable_key":"f8641d02-8413-5bd8-92e9-62ad49286e81:asa-graded-bulk-series","predicate":"limits","statement":"Using site-directed mutagenesis to prepare five further substitutions of Ser-530, the presence of amino acids with bulky side chains at position 530 inhibited cyclooxygenase activity and decreased the apparent affinity of the enzyme for arachidonate, consistent with the proposal that aspirin inhibits by placing a larger than normal side chain at position 530; residues 529 to 533 all proved important for the peroxidase as well as the cyclooxygenase activity, and Phe-529 in particular was critical for structure and catalysis.","claim_class":"mechanistic","status":"source_derived_draft","evidence_grade":"ungraded","direction":"negative","is_public":true,"mechanism_event_id":"2ac9a2e2-5497-5241-ba75-bb1e85308113","mechanism_event_label":"The bigger the group put at that position, the worse the enzyme binds its substrate.","subject":{"id":"7a465c55-930d-54cf-89b3-952b2c16f558","slug":"cox1-ser530","display_name":"Serine 530 of cyclooxygenase-1","entity_type_key":"protein_state"},"object":{"id":"e424fab2-cd9a-5b4c-b958-c3691c0e1b58","slug":"arachidonate-binding","display_name":"Binding of arachidonate in the cyclooxygenase active site","entity_type_key":"cellular_process"},"evidence_count":1,"mechanism_event":{"id":"2ac9a2e2-5497-5241-ba75-bb1e85308113","stable_key":"f8641d02-8413-5bd8-92e9-62ad49286e81:asa-graded-bulk-series-event","event_type":"biochemical_relationship","label":"The bigger the group put at that position, the worse the enzyme binds its substrate.","description":"Using site-directed mutagenesis to prepare five further substitutions of Ser-530, the presence of amino acids with bulky side chains at position 530 inhibited cyclooxygenase activity and decreased the apparent affinity of the enzyme for arachidonate, consistent with the proposal that aspirin inhibits by placing a larger than normal side chain at position 530; residues 529 to 533 all proved important for the peroxidase as well as the cyclooxygenase activity, and Phe-529 in particular was critical for structure and catalysis.","status":"provisional","compartment":null,"participants":[{"entity":{"id":"7afd60f8-544f-59a5-809f-c985bb5637eb","slug":"steric-blockade","display_name":"Steric blockade of a substrate channel by an introduced side chain","entity_type_key":"cellular_process"},"role":"mechanism","stoichiometry":null,"state_label":"","sequence_order":0,"notes":""},{"entity":{"id":"4f47dcac-4266-59eb-8c82-90777c27f21d","slug":"cyclooxygenase-activity","display_name":"The cyclooxygenase activity of prostaglandin endoperoxide synthase","entity_type_key":"cellular_process"},"role":"affected_activity","stoichiometry":null,"state_label":"","sequence_order":1,"notes":""},{"entity":{"id":"7a465c55-930d-54cf-89b3-952b2c16f558","slug":"cox1-ser530","display_name":"Serine 530 of cyclooxygenase-1","entity_type_key":"protein_state"},"role":"subject","stoichiometry":null,"state_label":"","sequence_order":2,"notes":""},{"entity":{"id":"e424fab2-cd9a-5b4c-b958-c3691c0e1b58","slug":"arachidonate-binding","display_name":"Binding of arachidonate in the cyclooxygenase active site","entity_type_key":"cellular_process"},"role":"target","stoichiometry":null,"state_label":"","sequence_order":3,"notes":""}]},"contexts":[{"dimension":"evidence_span","value_text":"{\"source_cache\": \"artifacts/aspirin-research/1601897.abstract.txt\", \"locator\": \"Indexed abstract; zero-based, end-exclusive Unicode character offsets\", \"file_sha256\": \"616cbc367eb38194b858872cbf91108baa3b1dec7aa039a40f69a29fa7157c2d\", \"start_char\": 0, \"end_char\": 1744, \"text_sha256\": \"616cbc367eb38194b858872cbf91108baa3b1dec7aa039a40f69a29fa7157c2d\"}","comparator":null,"unit":null,"notes":"","entity":null},{"dimension":"experimental_model","value_text":"Five further substitutions at Ser-530 and substitutions at adjoining residues, expressed transiently in cos-1 cells","comparator":null,"unit":null,"notes":"","entity":null},{"dimension":"exposure","value_text":"A graded series of side chains at position 530, plus substitutions at Phe-529, Leu-531, Lys-532 and Gly-533","comparator":null,"unit":null,"notes":"","entity":null},{"dimension":"limitations","value_text":"Tests the steric hypothesis by varying the bulk systematically rather than with one mutant. Transient expression in a heterologous cell line.","comparator":null,"unit":null,"notes":"","entity":null},{"dimension":"nutrient_topic","value_text":"Aspirin research collection; topical membership is not evidence of a direct clinical effect, and aspirin is recorded separately from salicylate, the metabolite it becomes.","comparator":null,"unit":null,"notes":"","entity":{"slug":"aspirin","display_name":"Aspirin / acetylsalicylic acid","entity_type_key":"drug"}},{"dimension":"organism","value_text":"Enzyme","comparator":null,"unit":null,"notes":"","entity":null},{"dimension":"plain_language","value_text":"The bigger the group put at that position, the worse the enzyme binds its substrate.","comparator":null,"unit":null,"notes":"","entity":null},{"dimension":"primary_references","value_text":"[asa-p1601897] Prostaglandin endoperoxide synthase. The aspirin acetylation region. (1992). https://pubmed.ncbi.nlm.nih.gov/1601897/ DOI: 10.1016/s0021-9258(19)49852-9","comparator":null,"unit":null,"notes":"","entity":null},{"dimension":"tissue_or_cell_type","value_text":"Recombinant prostaglandin endoperoxide H synthase-1","comparator":null,"unit":null,"notes":"","entity":null}],"evidence":[{"id":"44daf22c-8195-54df-9c39-4ef18b302817","evidence_kind":"source_excerpt","locator":"Lines 117-128","start_line":117,"end_line":128,"excerpt":"### asa-graded-bulk-series\nUsing site-directed mutagenesis to prepare five further substitutions of Ser-530, the presence of amino acids with bulky side chains at position 530 inhibited cyclooxygenase activity and decreased the apparent affinity of the enzyme for arachidonate, consistent with the proposal that aspirin inhibits by placing a larger than normal side chain at position 530; residues 529 to 533 all proved important for the peroxidase as well as the cyclooxygenase activity, and Phe-529 in particular was critical for structure and catalysis.\nCondition category: normal\nnutrient_topic: Aspirin research collection; topical membership is not evidence of a direct clinical effect, and aspirin is recorded separately from salicylate, the metabolite it becomes.\nplain_language: The bigger the group put at that position, the worse the enzyme binds its substrate.\norganism: Enzyme\ntissue_or_cell_type: Recombinant prostaglandin endoperoxide H synthase-1\nexperimental_model: Five further substitutions at Ser-530 and substitutions at adjoining residues, expressed transiently in cos-1 cells\nlimitations: Tests the steric hypothesis by varying the bulk systematically rather than with one mutant. Transient expression in a heterologous cell line.\nexposure: A graded series of side chains at position 530, plus substitutions at Phe-529, Leu-531, Lys-532 and Gly-533\nevidence_span: {\"source_cache\": \"artifacts/aspirin-research/1601897.abstract.txt\", \"locator\": \"Indexed abstract; zero-based, end-exclusive Unicode character offsets\", \"file_sha256\": \"616cbc367eb38194b858872cbf91108baa3b1dec7aa039a40f69a29fa7157c2d\", \"start_char\": 0, \"end_char\": 1744, \"text_sha256\": \"616cbc367eb38194b858872cbf91108baa3b1dec7aa039a40f69a29fa7157c2d\"}\n[asa-p1601897] Prostaglandin endoperoxide synthase. The aspirin acetylation region. (1992). https://pubmed.ncbi.nlm.nih.gov/1601897/ DOI: 10.1016/s0021-9258(19)49852-9","model_system":"Five further substitutions at Ser-530 and substitutions at adjoining residues, expressed transiently in cos-1 cells","directness":"author_interpretation","verification_status":"source_derived_draft","notes":"Exact curation-document quotation, not publisher quotation. Study references: [asa-p1601897] Prostaglandin endoperoxide synthase. The aspirin acetylation region. (1992). https://pubmed.ncbi.nlm.nih.gov/1601897/ DOI: 10.1016/s0021-9258(19)49852-9","relationship":"supports","weight":1.0,"link_notes":"","source":{"id":"712cf519-cd27-5a8b-9e2f-961a98d7a27e","stable_key":"import-f8641d02-8413-5bd8-92e9-62ad49286e81","title":"Aspirin: the serine it acetylates, the enzyme that acetylation creates, the dose that separates platelet from vessel wall, and the metabolite that is a different drug (2026-09-22)","document_type":"imported_text","citation_label":"AI-assisted literature curation; primary study URLs and scope retained in the document and extraction. Not publisher full text.","file_path":"","sha256":"93dd4343051c131e40d4fc3c95a978cd65478c13dcdb3f2d42507c49b09cb647","revision_id":"694e8ee7-49ca-5e6c-b58b-8ba8f82c2ad8","review_status":"unverified_draft","notes":""}}],"relations":[],"conflicts":[],"corrections":[],"research":null}