title: Prostaglandin H synthase-2-catalyzed oxygenation of 2-arachidonoylglycerol is more sensitive to peroxide tone than oxygenation of arachidonic acid.
abstract: The endocannabinoid, 2-arachidonoylglycerol (2-AG), is a selective substrate for the inducible isoform of prostaglandin H synthase (PGHS), PGHS-2. Its turnover leads to the formation of glyceryl esters of prostaglandins (PG-Gs), a subset of which elicits agonism at unique, as yet unidentified, receptors. The k(cat)/K(m) values for oxygenation of arachidonic acid (AA) and 2-AG by PGHS-2 are very similar, but the sensitivities of the two substrates to peroxide-dependent activation have not been compared. 15-Hydroperoxy derivatives of AA and 2-AG were found to be comparable in their ability to serve as substrates for the peroxidase activities of PGHS-2, PGHS-1, and glutathione peroxidase (GPx). They also were comparable in the activation of AA oxygenation by cyanide-inhibited PGHS-2. However, oxygenation of 2-AG was significantly suppressed relative to AA by the presence of GPx and GSH. Furthermore, 2-AG oxygenation by peroxidase-deficient H388YmPGHS-2 was much less efficient than AA oxygenation. Wild-type rates of 2-AG oxygenation were restored by treatment of H388YmPGHS-2 with hydroperoxide derivatives of AA or 2-AG. RNAi silencing of phospholipid hydroperoxide-specific GPx (GPx4) in NIH/3T3 cells led to increases in cellular peroxidation and in the levels of the isoprostane product, 8-epi-PGF(2α). GPx4 silencing led to 2-4-fold increases in PG-G formation but no change in PG formation. Thus, cellular peroxide tone may be an important determinant of the extent of endocannabinoid oxygenation by PGHS-2.
authors.affiliation: A. B. Hancock Jr. Memorial Laboratory for Cancer Research, Department of Biochemistry, Vanderbilt Institute of Chemical Biology, Center in Molecular Toxicology, Vanderbilt University School of Medicine, Nashville, Tennessee 37232-0146, USA.
authors.forename: Joel
authors.initials: J
authors.lastname: Musee
authors.forename: Lawrence J
authors.initials: LJ
authors.lastname: Marnett
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D016718:Arachidonic Acid / Q000737:chemistry* / Q000378:metabolism; D001095:Arachidonic Acids / Q000737:chemistry* / Q000378:metabolism; D052160:Benzothiazoles / Q000737:chemistry; D055162:Biocatalysis; D002417:Cattle; D002863:Chromogenic Compounds / Q000737:chemistry; D051545:Cyclooxygenase 1 / Q000737:chemistry; D051546:Cyclooxygenase 2 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D063388:Endocannabinoids / Q000737:chemistry* / Q000378:metabolism; D004789:Enzyme Activation; D055785:Gene Knockdown Techniques; D005979:Glutathione Peroxidase / Q000235:genetics / Q000378:metabolism; D005989:Glycerides / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D051379:Mice; D041681:NIH 3T3 Cells; D010084:Oxidation-Reduction; D018384:Oxidative Stress; D010545:Peroxides / Q000737:chemistry* / Q000378:metabolism; D000080662:Phospholipid Hydroperoxide Glutathione Peroxidase; D011453:Prostaglandins / Q000096:biosynthesis; D034622:RNA Interference; D013451:Sulfonic Acids / Q000737:chemistry; D000094765:Glutathione Peroxidase GPX1
chemical_list: D001095:Arachidonic Acids; D052160:Benzothiazoles; D002863:Chromogenic Compounds; D063388:Endocannabinoids; D005989:Glycerides; D010545:Peroxides; D011453:Prostaglandins; D013451:Sulfonic Acids; D016718:Arachidonic Acid; C002502:2,2'-azino-di-(3-ethylbenzothiazoline)-6-sulfonic acid; C094503:glyceryl 2-arachidonate; D000080662:Phospholipid Hydroperoxide Glutathione Peroxidase; D005979:Glutathione Peroxidase; D051545:Cyclooxygenase 1; D051546:Cyclooxygenase 2; C496511:PTGS1 protein, human; D000094765:Glutathione Peroxidase GPX1