Pre-existent asymmetry in the human cyclooxygenase-2 sequence homodimer

  • Posted on: 5 November 2014
  • By: fcoldren
TitlePre-existent asymmetry in the human cyclooxygenase-2 sequence homodimer
Publication TypeJournal Article
Year of Publication2013
AuthorsDong L, Sharma NP, Jurban BJ, Smith WL
JournalJ Biol Chem
Date Published2013 Oct 4
KeywordsAcetylation, Amino Acid Sequence, Amino Acid Substitution, Anti-Inflammatory Agents, Non-Steroidal, Arachidonic Acid, Aspirin, Cyclooxygenase 2, Cyclooxygenase Inhibitors, Flurbiprofen, Guanidine, Heme, Humans, Indomethacin, Kinetics, Models, Biological, Mutant Proteins, Mutation, Naproxen, Oxygen, Palmitic Acid, Peroxidase, Protein Multimerization, Pyrazoles, Substrate Specificity, Sulfonamides

Prostaglandin endoperoxide H synthase-2 (PGHS-2), also known as cyclooxygenase-2 (COX-2), is a sequence homodimer. However, the enzyme exhibits half-site heme and inhibitor binding and functions as a conformational heterodimer having a catalytic subunit (Ecat) with heme bound and an allosteric subunit (Eallo) lacking heme. Some recombinant heterodimers composed of a COX-deficient mutant subunit and a native subunit (i.e. Mutant/Native PGHS-2) have COX activities similar to native PGHS-2. This suggests that the presence of heme plus substrate leads to the subunits becoming lodged in a semi-stable Eallo-mutant/Ecat-Native∼heme form during catalysis. We examined this concept using human PGHS-2 dimers composed of combinations of Y385F, R120Q, R120A, and S530A mutant or native subunits. With some heterodimers (e.g. Y385F/Native PGHS-2), heme binds with significantly higher affinity to the native subunit. This correlates with near native COX activity for the heterodimer. With other heterodimers (e.g. S530A/Native PGHS-2), heme binds with similar affinities to both subunits, and the COX activity approximates that expected for an enzyme in which each monomer contributes equally to the net COX activity. With or without heme, aspirin acetylates one-half of the subunits of the native PGHS-2 dimer, the Ecat subunits. Subunits having an S530A mutation are refractory to acetylation. Curiously, aspirin acetylates only one-quarter of the monomers of S530A/Native PGHS-2 with or without heme. This implies that there are comparable amounts of two noninterchangeable species of apoenzymes, Eallo-S530A/Ecat-Native and Eallo-Native/Ecat-S530A. These results suggest that native PGHS-2 assumes a reasonably stable, asymmetric Eallo/Ecat form during its folding and processing.

Alternate JournalJ. Biol. Chem.
PubMed ID23955344
PubMed Central IDPMC3789963
Grant ListCA130810 / CA / NCI NIH HHS / United States
GM68848 / GM / NIGMS NIH HHS / United States
HL117798 / HL / NHLBI NIH HHS / United States
P50 CA130810 / CA / NCI NIH HHS / United States
R01 GM068848 / GM / NIGMS NIH HHS / United States