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High-level expression of active human cyclooxygenase-2 in insect cells
W A Cromlish1, P Payette, S A Culp
1Department of Biochemistry and Molecular Biology, Merck Frosst Center for Therapeutic Research, Merck Frosst Canada, Inc., Pointe-Claire-Dorval, Quebec, Canada.
Archives of Biochemistry and Biophysics
|October 1, 1994
Summary
Researchers successfully expressed active human cyclooxygenase-2 (Cox-2) in insect cells. The insect-derived enzyme exhibited comparable activity and drug inhibition profiles to native Cox-2, validating this expression system.
Area of Science:
- Biochemistry
- Molecular Biology
- Enzymology
Background:
- Cyclooxygenase-2 (Cox-2) is a key enzyme in inflammation and pain pathways.
- Efficient expression systems are crucial for studying Cox-2 function and developing anti-inflammatory drugs.
Purpose of the Study:
- To express active human cyclooxygenase-2 (Cox-2) in insect cells using a recombinant baculovirus system.
- To characterize the enzymatic activity, glycosylation, and drug inhibition profile of the expressed Cox-2.
Main Methods:
- Recombinant baculovirus was used to express human Cox-2 in insect cells.
- Enzyme activity was measured in microsomes from infected cells.
- Glycosylation patterns and inhibition by nonsteroidal anti-inflammatory drugs (NSAIDs) were analyzed.
Main Results:
- High levels of active human Cox-2 were expressed in insect cells.
- The specific activity of insect-derived Cox-2 was comparable to ovine placenta Cox-2.
- The expressed enzyme showed characteristic aspirin-induced 15-hydroxyeicosa-tetraenoic acid production and typical NSAID inhibition.
Conclusions:
- Insect cells provide a viable system for expressing functional human Cox-2.
- The expressed enzyme retains key biochemical and pharmacological properties, useful for drug discovery and research.