Aspirin-acetylated cyclooxygenase-1

evidence_span

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experimental_model

X-ray crystal structure at 3.4 angstrom of prostaglandin H2 synthase-1 inactivated by a potent aspirin analogue

exposure

2-bromoacetoxy-benzoic acid acetylation, with salicylic acid soaked into the structure

limitations

Direct structural evidence rather than inference from mutants, though at modest resolution and using an analogue rather than aspirin itself.

nutrient_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. · Aspirin / acetylsalicylic acid

organism

Enzyme

plain_language

The picture confirms it: the acetyl group simply plugs the tunnel the fatty acid has to travel down.

primary_references

[asa-p7552725] The structural basis of aspirin activity inferred from the crystal structure of inactivated prostaglandin H2 synthase. (1995). https://pubmed.ncbi.nlm.nih.gov/7552725/ DOI: 10.1038/nsb0895-637

tissue_or_cell_type

Crystallised prostaglandin H2 synthase-1

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) · lines 130–141

AI-assisted literature curation; primary study URLs and scope retained in the document and extraction. Not publisher full text. · supports · X-ray crystal structure at 3.4 angstrom of prostaglandin H2 synthase-1 inactivated by a potent aspirin analogue · source_derived_draft · unverified_draft

### asa-structure-shows-blockage The 3.4 angstrom X-ray crystal structure of prostaglandin H2 synthase isoform-1 inactivated by the potent aspirin analogue 2-bromoacetoxy-benzoic acid shows that acetylation abolishes cyclooxygenase activity by steric blockage of the active-site channel and not through a large conformational change, with two rotameric states of the acetyl-serine side chain blocking the channel to different extents, a result which may explain the dissimilar effects of aspirin on the two isoforms. Condition category: normal nutrient_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. plain_language: The picture confirms it: the acetyl group simply plugs the tunnel the fatty acid has to travel down. organism: Enzyme tissue_or_cell_type: Crystallised prostaglandin H2 synthase-1 experimental_model: X-ray crystal structure at 3.4 angstrom of prostaglandin H2 synthase-1 inactivated by a potent aspirin analogue limitations: Direct structural evidence rather than inference from mutants, though at modest resolution and using an analogue rather than aspirin itself. exposure: 2-bromoacetoxy-benzoic acid acetylation, with salicylic acid soaked into the structure evidence_span: {"source_cache": "artifacts/aspirin-research/7552725.abstract.txt", "locator": "Indexed abstract; zero-based, end-exclusive Unicode character offsets", "file_sha256": "dd61431f20eb312313942bc8a89f970d5d2e33600b32abe692f20d9d5c57d7b1", "start_char": 0, "end_char": 751, "text_sha256": "dd61431f20eb312313942bc8a89f970d5d2e33600b32abe692f20d9d5c57d7b1"} [asa-p7552725] The structural basis of aspirin activity inferred from the crystal structure of inactivated prostaglandin H2 synthase. (1995). https://pubmed.ncbi.nlm.nih.gov/7552725/ DOI: 10.1038/nsb0895-637

evidence_span

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experimental_model

Site-directed mutagenesis of sheep prostaglandin endoperoxide synthase expressed in Cos-1 cells

exposure

Serine 530 replaced with alanine, compared with the native enzyme against aspirin and reversible inhibitors

limitations

The decisive experiment for the mechanism: it separates what the serine does for catalysis from what acetylating it does. Recombinant sheep enzyme, not human enzyme in a tissue.

nutrient_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. · Aspirin / acetylsalicylic acid

organism

Sheep and mouse enzyme

plain_language

The acetyl group works by being in the way, not by removing something the enzyme needed.

primary_references

[asa-p2108169] The aspirin and heme-binding sites of ovine and murine prostaglandin endoperoxide synthases. (1990). https://pubmed.ncbi.nlm.nih.gov/2108169/ DOI: 10.1016/s0021-9258(19)34105-5

tissue_or_cell_type

Recombinant enzyme in Cos-1 cells

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) · lines 91–102

AI-assisted literature curation; primary study URLs and scope retained in the document and extraction. Not publisher full text. · supports · Site-directed mutagenesis of sheep prostaglandin endoperoxide synthase expressed in Cos-1 cells · source_derived_draft · unverified_draft

### asa-acetylation-is-steric Acetylation of Ser-530 of sheep prostaglandin endoperoxide synthase by aspirin causes irreversible inactivation of the cyclooxygenase activity of the enzyme, and since the serine hydroxyl is not required for catalysis the acetylation apparently introduces a bulky side chain at position 530 which interferes with arachidonate binding; a sequence of 35 residues with Ser-530 at the midpoint was identical in the sheep and mouse enzymes. Condition category: normal nutrient_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. plain_language: The acetyl group works by being in the way, not by removing something the enzyme needed. organism: Sheep and mouse enzyme tissue_or_cell_type: Recombinant enzyme in Cos-1 cells experimental_model: Site-directed mutagenesis of sheep prostaglandin endoperoxide synthase expressed in Cos-1 cells limitations: The decisive experiment for the mechanism: it separates what the serine does for catalysis from what acetylating it does. Recombinant sheep enzyme, not human enzyme in a tissue. exposure: Serine 530 replaced with alanine, compared with the native enzyme against aspirin and reversible inhibitors evidence_span: {"source_cache": "artifacts/aspirin-research/2108169.abstract.txt", "locator": "Indexed abstract; zero-based, end-exclusive Unicode character offsets", "file_sha256": "c7b11508521a3854b432f3836de6e92ba9863c8edb244d747bb1c37634c0bb50", "start_char": 0, "end_char": 2076, "text_sha256": "c7b11508521a3854b432f3836de6e92ba9863c8edb244d747bb1c37634c0bb50"} [asa-p2108169] The aspirin and heme-binding sites of ovine and murine prostaglandin endoperoxide synthases. (1990). https://pubmed.ncbi.nlm.nih.gov/2108169/ DOI: 10.1016/s0021-9258(19)34105-5

evidence_span

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experimental_model

Human cyclooxygenase-1 and -2 expressed in cos-1 cells treated with aspirin and with carbon-14 labelled aspirin, with a mutant series at Ser-516

exposure

Aspirin acetylation, with S516A, S516N, S516Q and S516M substitutions

limitations

Identifies the site by direct labelling and explains the isoform difference by active site volume. Recombinant enzyme in a heterologous cell line.

nutrient_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. · Aspirin / acetylsalicylic acid

organism

Human enzyme

plain_language

Acetylating the second enzyme does not switch it off; it turns it into a machine that makes something else.

primary_references

[asa-p8175750] Acetylation of human prostaglandin endoperoxide synthase-2 (cyclooxygenase-2) by aspirin. (1994). https://pubmed.ncbi.nlm.nih.gov/8175750/ DOI: 10.1016/s0021-9258(17)36820-5

tissue_or_cell_type

Recombinant cyclooxygenase

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) · lines 208–219

AI-assisted literature curation; primary study URLs and scope retained in the document and extraction. Not publisher full text. · supports · Human cyclooxygenase-1 and -2 expressed in cos-1 cells treated with aspirin and with carbon-14 labelled aspirin, with a mutant series at Ser-516 · source_derived_draft · unverified_draft

### asa-cox2-becomes-a-lipoxygenase Aspirin treatment of human cyclooxygenase-1 caused time-dependent inactivation of oxygenase activity, whereas aspirin treatment of cyclooxygenase-2 produced an enzyme which retained oxygenase activity but formed exclusively 15-hydroxyeicosatetraenoic acid instead of prostaglandin H2, exclusively of the 15R configuration, with Km values for arachidonate of native and aspirin-treated cyclooxygenase-2 about the same, suggesting arachidonate binds to both in a similar manner. Condition category: normal nutrient_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. plain_language: Acetylating the second enzyme does not switch it off; it turns it into a machine that makes something else. organism: Human enzyme tissue_or_cell_type: Recombinant cyclooxygenase experimental_model: Human cyclooxygenase-1 and -2 expressed in cos-1 cells treated with aspirin and with carbon-14 labelled aspirin, with a mutant series at Ser-516 limitations: Identifies the site by direct labelling and explains the isoform difference by active site volume. Recombinant enzyme in a heterologous cell line. exposure: Aspirin acetylation, with S516A, S516N, S516Q and S516M substitutions evidence_span: {"source_cache": "artifacts/aspirin-research/8175750.abstract.txt", "locator": "Indexed abstract; zero-based, end-exclusive Unicode character offsets", "file_sha256": "9ba85af90c62b157cf53032375ca703e8b78eff6ddd22ad627bbd7e65965d8e9", "start_char": 0, "end_char": 1786, "text_sha256": "9ba85af90c62b157cf53032375ca703e8b78eff6ddd22ad627bbd7e65965d8e9"} [asa-p8175750] Acetylation of human prostaglandin endoperoxide synthase-2 (cyclooxygenase-2) by aspirin. (1994). https://pubmed.ncbi.nlm.nih.gov/8175750/ DOI: 10.1016/s0021-9258(17)36820-5

evidence_span

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experimental_model

Site-directed mutagenesis of mouse COX-2 residues predicted from comparison of the two isoform crystal structures

exposure

Val-434, Arg-513, Val-523, Val-228 and Gly-533 mutated towards their cyclooxygenase-1 equivalents, then acetylated

limitations

Explains the isoform difference residue by residue. Murine enzyme, and the L-shaped substrate configuration is an inference from the mutant product profiles.

nutrient_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. · Aspirin / acetylsalicylic acid

organism

Mouse enzyme

plain_language

Three amino acids are the whole difference between an enzyme aspirin kills and one aspirin repurposes.

primary_references

[asa-p10692466] Spatial requirements for 15-(R)-hydroxy-5Z,8Z,11Z, 13E-eicosatetraenoic acid synthesis within the cyclooxygenase active site of murine COX-2. Why acetylated COX-1 does not synthesize 15-(R)-hete. (2000). https://pubmed.ncbi.nlm.nih.gov/10692466/ DOI: 10.1074/jbc.275.9.6586

tissue_or_cell_type

Cyclooxygenase-2

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) · lines 247–258

AI-assisted literature curation; primary study URLs and scope retained in the document and extraction. Not publisher full text. · supports · Site-directed mutagenesis of mouse COX-2 residues predicted from comparison of the two isoform crystal structures · source_derived_draft · unverified_draft

### asa-three-residues-decide Acetylated cyclooxygenase-2 is essentially a lipoxygenase making 15(R)- and 11(R)-hydroxyeicosatetraenoic acid whereas acetylated cyclooxygenase-1 is unable to oxidise arachidonic acid to any products, and site-directed mutagenesis of Val-434, Arg-513 and Val-523 in mouse cyclooxygenase-2 to their cyclooxygenase-1 equivalents abrogated 11- and 15-HETE production after aspirin treatment, confirming these residues as the major isoform selectivity determinants; V228F and G533A produced wild-type-like profiles but on acetylation neither could make HETE products, suggesting arachidonic acid orientates in an L-shaped binding configuration. Condition category: normal nutrient_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. plain_language: Three amino acids are the whole difference between an enzyme aspirin kills and one aspirin repurposes. organism: Mouse enzyme tissue_or_cell_type: Cyclooxygenase-2 experimental_model: Site-directed mutagenesis of mouse COX-2 residues predicted from comparison of the two isoform crystal structures limitations: Explains the isoform difference residue by residue. Murine enzyme, and the L-shaped substrate configuration is an inference from the mutant product profiles. exposure: Val-434, Arg-513, Val-523, Val-228 and Gly-533 mutated towards their cyclooxygenase-1 equivalents, then acetylated evidence_span: {"source_cache": "artifacts/aspirin-research/10692466.abstract.txt", "locator": "Indexed abstract; zero-based, end-exclusive Unicode character offsets", "file_sha256": "9da1ede1fbf95d6f42d4bb15dc08b5d1869549a7cd94cbfa9da9f9865fd780ff", "start_char": 0, "end_char": 2055, "text_sha256": "9da1ede1fbf95d6f42d4bb15dc08b5d1869549a7cd94cbfa9da9f9865fd780ff"} [asa-p10692466] Spatial requirements for 15-(R)-hydroxy-5Z,8Z,11Z, 13E-eicosatetraenoic acid synthesis within the cyclooxygenase active site of murine COX-2. Why acetylated COX-1 does not synthesize 15-(R)-hete. (2000). https://pubmed.ncbi.nlm.nih.gov/10692466/ DOI: 10.1074/jbc.275.9.6586