Abstract
BackgroundRecent studies showed that some of the dietary bioflavonoids can strongly stimulate the catalytic activity of cyclooxygenase (COX) I and II in vitro and in vivo, presumably by facilitating enzyme re-activation. In this study, we sought to understand the structural basis of COX activation by these dietary compounds.Methodology/Principal FindingsA combination of molecular modeling studies, biochemical analysis and site-directed mutagenesis assay was used as research tools. Three-dimensional quantitative structure-activity relationship analysis (QSAR/CoMFA) predicted that the ability of bioflavonoids to activate COX I and II depends heavily on their B-ring structure, a moiety known to be associated with strong antioxidant ability. Using the homology modeling and docking approaches, we identified the peroxidase active site of COX I and II as the binding site for bioflavonoids. Upon binding to this site, bioflavonoid can directly interact with hematin of the COX enzyme and facilitate the electron transfer from bioflavonoid to hematin. The docking results were verified by biochemical analysis, which reveals that when the cyclooxygenase activity of COXs is inhibited by covalent modification, myricetin can still stimulate the conversion of PGG2 to PGE2, a reaction selectively catalyzed by the peroxidase activity. Using the site-directed mutagenesis analysis, we confirmed that Q189 at the peroxidase site of COX II is essential for bioflavonoids to bind and re-activate its catalytic activity.Conclusions/SignificanceThese findings provide the structural basis for bioflavonoids to function as high-affinity reducing co-substrates of COXs through binding to the peroxidase active site, facilitating electron transfer and enzyme re-activation.
Highlights
Cyclooxygenase (COX) I and II catalyze the metabolism of arachidonic acid (AA), resulting in the formation of prostaglandins (PGs), thromboxanes, and hydroxyeicosateraenoic acids (HETEs) [1,2,3,4], which exert an array of important biological actions in the body [5,6,7]
To probe the structural determinants of various bioflavonoids for activating COX I and II, we developed the 3-D Quantitative structure-activity relationship (QSAR)/ CoMFA models by using the experimental data obtained from 9 representative bioflavonoids and flavone
We conducted computational molecular modeling study and biochemical analysis to probe the mechanism of COX activation by dietary bioflavonoids
Summary
Cyclooxygenase (COX) I and II catalyze the metabolism of arachidonic acid (AA), resulting in the formation of prostaglandins (PGs), thromboxanes, and hydroxyeicosateraenoic acids (HETEs) [1,2,3,4], which exert an array of important biological actions in the body [5,6,7]. Whereas COX I is a constitutively-expressed enzyme in most tissues and primarily functions as a house-keeping enzyme, COX II is a highlyinducible enzyme (such as in the presence of various mitogens) and plays a critical role in mediating inflammation [11] Both COX I and II have two catalytic activities that are functionally coupled: one is to catalyze the cyclooxygenase reaction that converts AA to prostaglandin G2 (PGG2), and the other one is to catalyzes the peroxidase reaction that reduces PGG2 to prostaglandin H2 (PGH2). A branched-chain model has been proposed to explain the mechanism of those two reactions catalyzed by COX I and II [12] Based on this model, a peroxide (such as PGG2) is thought to initiate the peroxidase reaction by abstracting two electrons from hematin in the peroxidase active site, yielding Compound I, a protoporphyrin IX (PPIX) radical cation with an oxyferry group (Fe4+ = O). We sought to understand the structural basis of COX activation by these dietary compounds
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