Arachidonate cyclo-oxygenase (prostaglandin synthetase; prostaglandin endoperoxide synthetase; EC 1.14.99.1) was purified from sheep platelets. The purification procedure involved hydrophobic column chromatography using either Ibuprofen-Sepharose, phenyl-Sepharose or arachidic acid-Sepharose as the first step followed by metal-chelate Sepharose and haemin-Sepharose affinity chromatography. The purified enzyme (Mr approximately 65,000) was homogeneous as observed by SDS/polyacrylamide-gel electrophoresis and silver staining. The enzyme was a glycoprotein with mannose as the neutral sugar. Haemin or haemoglobin was essential for activity. The purified enzyme could bind haemin exhibiting a characteristic absorption maximum at 410 nm. The enzyme after metal-chelate column chromatography could undergo acetylation by [acetyl-3H]aspirin. The labelled acetylated enzyme could not bind to haemin-Sepharose, presumably due to acetylation of a serine residue involved in the binding to haemin. The acetylated enzyme also failed to show its characteristic absorption maximum at 410 nm when allowed to bind haemin.
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October 1986
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Research Article|
October 15 1986
Purification and characterization of sheep platelet cyclo-oxygenase Acetylation by aspirin prevents haemin binding to the enzyme
Publisher: Portland Press Ltd
Online ISSN: 1470-8728
Print ISSN: 0264-6021
© 1986 London: The Biochemical Society
1986
Biochem J (1986) 239 (2): 371–377.
Citation
R Boopathy, A S Balasubramanian; Purification and characterization of sheep platelet cyclo-oxygenase Acetylation by aspirin prevents haemin binding to the enzyme. Biochem J 15 October 1986; 239 (2): 371–377. doi: https://doi.org/10.1042/bj2390371
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