Intracellular Erythrocyte Platelet-activating Factor Acetylhydrolase I Inactivates Aspirin in Blood

• Published in: The Journal of biological chemistry Vol. 286; no. 40; pp. 34820 - 34829
• Main Authors: Zhou, Gang, Marathe, Gopal K., Willard, Belinda, McIntyre, Thomas M.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 07.10.2011; American Society for Biochemistry and Molecular Biology
• Subjects: 1-Alkyl-2-acetylglycerophosphocholine Esterase - blood; 1-Alkyl-2-acetylglycerophosphocholine Esterase - genetics; 1-Alkyl-2-acetylglycerophosphocholine Esterase - metabolism; Amino Acid Sequence; Aspirin; Aspirin - pharmacology; Blood; Carboxylic Ester Hydrolases - metabolism; Chromatography - methods; Chromatography, Gel; Cyclooxygenase (COX) Pathway; Drug Metabolism; Drug Resistance; Eicosanoid Regulation; Enzyme Purification; Enzymology; Erythrocyte; Erythrocytes - cytology; Erythrocytes - drug effects; Humans; Hydrolysis; Kinetics; Mass Spectrometry - methods; Microtubule-Associated Proteins - blood; Microtubule-Associated Proteins - genetics; Models, Statistical; Molecular Sequence Data; Trypsin - chemistry; Enzyme Purification; Erythrocyte; Aspirin; Drug Metabolism; Cyclooxygenase (COX) Pathway; Blood; Drug Resistance; Eicosanoid Regulation
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M111.267161

Aspirin (acetylsalicylic acid) prophylaxis suppresses major adverse cardiovascular events, but its rapid turnover limits inhibition of platelet cyclooxygenase activity and thrombosis. Despite its importance, the identity of the enzyme(s) that hydrolyzes the acetyl residue of circulating aspirin, which must be an existing enzyme, remains unknown. We find that circulating aspirin was extensively hydrolyzed within erythrocytes, and chromatography indicated these cells contained a single hydrolytic activity. Purification by over 1400-fold and sequencing identified the PAFAH1B2 and PAFAH1B3 subunits of type I platelet-activating factor (PAF) acetylhydrolase, a phospholipase A2 with selectivity for acetyl residues of PAF, as a candidate for aspirin acetylhydrolase. Western blotting showed that catalytic PAFAH1B2 and PAFAH1B3 subunits of the type I enzyme co-migrated with purified erythrocyte aspirin hydrolytic activity. Recombinant PAFAH1B2, but not its family member plasma PAF acetylhydrolase, hydrolyzed aspirin, and PAF competitively inhibited aspirin hydrolysis by purified or recombinant erythrocyte enzymes. Aspirin was hydrolyzed by HEK cells transfected with PAFAH1B2 or PAFAH1B3, and the competitive type I PAF acetylhydrolase inhibitor NaF reduced erythrocyte hydrolysis of aspirin. Exposing aspirin to erythrocytes blocked its ability to inhibit thromboxane A2 synthesis and platelet aggregation. Not all individuals or populations are equally protected by aspirin prophylaxis, the phenomenon of aspirin resistance, and erythrocyte hydrolysis of aspirin varied 3-fold among individuals, which correlated with PAFAH1B2 and not PAFAH1B3. We conclude that intracellular type I PAF acetylhydrolase is the major aspirin hydrolase of human blood. Background: Aspirin circulates transiently in blood, but the identity of the enzyme(s) that hydrolyzes its acetyl residue remains unknown. Results: Purification, mass spectrometry, and overexpression identified erythrocyte type I PAF acetylhydrolase as aspirin hydrolase. Conclusion: Aspirin is primarily hydrolyzed within erythrocytes by PAF acetylhydrolase. Significance: PAF acetylhydrolase and aspirin hydrolysis varies among individuals to modulate the effectiveness of aspirin.