Title : Crystal Structure of Aspirin-Acetylated
Human Cyclooxygenase-2 : Insight into the Formation of Products with Reversed Stereochemistry
Abstract :
- Aspirin and other nonsteroidal anti-inflammatory drugs target the cyclooxygenase enzymes ( COX-1 and COX-2 ) to block the formation of prostaglandins
- Aspirin is unique in that it covalently modifies each enzyme by acetylating Ser-530 within the cyclooxygenase active site
- Acetylation of COX-1 leads to complete loss of activity, while acetylation of COX-2 results in the generation of the monooxygenated product 15(R)-hydroxyeicosatetraenoic acid (15R-HETE)
- Ser-530 has also been shown to influence the stereochemistry for the addition of oxygen to the prostaglandin product
- We determined the crystal structures of S530T murine (mu) COX-2 , aspirin-acetylated human (hu) COX-2, and hu COX-2 in complex with salicylate to 1.9, 2.0, and 2.4 Å, respectively
- The structures reveal that (1) the acetylated Ser-530 completely blocks access to the hydrophobic groove, (2) the observed binding pose of salicylate is reflective of the enzyme-inhibitor complex prior to acetylation, and (3) the observed Thr-530 rotamer in the S530T mu COX-2 crystal structure does not impede access to the hydrophobic groove
- On the basis of these structural observations, along with functional analysis of the S530T/G533V double mutant, we propose a working hypothesis for the generation of 15R-HETE by aspirin-acetylated COX-2
- We also observe differential acetylation of COX-2 purified in various detergent systems and nanodiscs, indicating that detergent and lipid binding within the membrane-binding domain of the enzyme alters the rate of the acetylation reaction in vitro