Analysis of Cyclooxygenase-Substrate Interactions Using Stereospecificallylabeled Arachidonic Acids
The two mammalian cyclooxygenase isoforms, COX-1 and COX-2, catalyze an identical reaction in the formation of prostaglandin H2 from arachidonic acid (Smith et al., 1996). The initial abstraction of the pro-S hydrogen at carbon C-13 is followed by oxygenation in the 11R position, formation of the endoperoxide and the five-membered prostaglandin ring, and, finally, a second oxygenation at C-15 to yield prostaglandin H2 (Hamberg and Samuelsson, 1967a).
As with all cyclooxygenases and lipoxygenases investigated so far, this mechanism follows the so-called antarafacial rule of hydrogen abstraction and oxygen insertion: both events occur on opposite sites of the planar cis,cis-pentadiene system of arachidonic acid (Hamberg and Samuelsson, 1967a; 1967b). COX-1 and COX-2, however, are different in their response to treatment with the non-steroidal anti-inflammatory drug, aspirin. In both enzymes, aspirin acetylates a critical serine residue in the active site channel. This leads to a complete block of oxygenase activity in COX-1, but to a new oxygenase specificity in COX-2, namely, formation of 15R-hydroxyeicosatetraenoic acid (15R-HETE) as the sole enzymatic product (Holtzman et al., 1992; Meade et al., 1993; Lecomte et al., 1994).
KeywordsHPLC Hydroxyl Catalysis Aspirin Serine
Unable to display preview. Download preview PDF.
- 4.Kurumbail, R.G., Stevens, A.M., Gierse, J.K., McDonald, J.J., Stegeman, R.A., Pak, J.Y., Gildehaus, J.M., Miyashiro, J.M., Penning, T.D., Seibert, K., Isakson, P.C., and Stallings, W.C., 1996, Structural basis for selective inhibition of cyclooxygenase-2 by anti-inflammatory agentsNature384:644.PubMedCrossRefGoogle Scholar
- 5.Laneuville, O., Breuer, D.K., Xu, N., Huang, Z.H., Gage, D.A., Watson, J.T., Lagarde, M., DeWitt, D.L., and Smith, W.L., 1995, Fatty acid substrate specificities of human prostaglandin-endoperoxide H synthase-1 and -2. Formation of t2-hydroxy-(9Z, I 3E/Z, 15Z)-octadecatrienoic acids from alphalinolenic acidJ. Biol. Chem.270:19330.PubMedCrossRefGoogle Scholar
- 9.Rowlinson, S.W., Crews, B.C., Lanzo, C.A., and Marnett, L.J., 1999, The binding of arachidonic acid in the cyclo-oxygenase active site of mouse prostaglandin endoperoxide synthase-2 (COX-2). A putative L-shaped binding conformation utilizing the top channel regionJ. Biol. Chem.274:23305.PubMedCrossRefGoogle Scholar