Journal of lipid mediators最新文献

Leukotriene (LT) A4 hydrolase catalyzes enzymatic hydration of LTA4 to biologically active substance, LTB4. Biochemical and immuno-histochemical studies have shown that this enzyme is ubiquitously distributed in various cells and tissues. A sequence domain of LTA4 hydrolase was found to be homologous to those of several zinc metalloproteases. Both native and recombinant enzymes were shown to possess equimolar zinc ion and aminopeptidase activity. To examine the molecular mechanism of this enzyme reaction, site-directed mutagenesis experiments were carried out. Single amino acid substitutions at Glu-297 revealed a distinction of two enzyme activities, and suggest that the glutamic acid residue at 297 is essential for aminopeptidase, while the side chain of Glu or Gln is required for LTA4 hydrolase activity. The loss of two enzyme activities in a mutant E319K confirmed the proposal that the presence of a zinc ion in the enzyme is required for both enzyme activities.

The receptors involved in the ocular hypotensive activity of prostaglandins (PG) E2 and F2 alpha in dogs and monkeys was investigated by examining the effects of putative receptor selective agonists on intraocular pressure. A diverse variety of receptor selective agonists lowered intraocular pressure in these species. Thus, FP-receptor agonists (17-phenyl PGF2 alpha, fluprostenol), agonists with potent activity at the EP3 receptor (MB 28767, sulprostone) and a prostanoid with activity at the EP2 receptor (11-deoxy PGE1) were all potent ocular hypotensives when administered as a single dose to dogs and monkeys or b.i.d. for 5 days in monkeys. These findings were regarded as surprising and prompted us to re-examine some aspects of the current classification for prostanoid receptors. At present certain receptor subtypes, notably EP2, EP3 and FP receptors, are defined only according to the potency rank order for agonists. In these studies, we employed radioligand binding studies to determine the degree of competition between prostanoid agonists claimed to be selective on the basis of functional assays. Competition studies with a diverse variety of prostanoids at the binding site for PGE2 and sulprostone on the myometrial plasma membrane prepared from the rat uterus were consistent with the presence of an EP3 receptor. Thus, EP3-receptor agonists (MB 28767 and sulprostone) potently inhibited PGE2 and sulprostone binding, whereas FP agonists (17-phenyl PGF2 alpha, fluprostenol), a DP agonist (BW 245C), an EP1 antagonist (AH 6809), an EP2 agonist (AH 13205) and TP-receptor ligands (BM 13505, I-BOP) afforded little or no inhibition.(ABSTRACT TRUNCATED AT 250 WORDS)

A gene subfamily of cytochromes P450 with catalytic activity toward various eicosanoid substrates has been studied with a variety of techniques in this laboratory, including purification and characterization, localization at the tissue and subcellular levels, physiological function, and cloning and expression in prokaryotic and eukaryotic systems. This paper reports experiments directed toward determining the function of the cytochrome P4504A metabolite, 20-hydroxyarachidonic acid (20-hydroxyeicosatetraenoic acid; 20-HETE), in cellular ion flux, immunohistochemical localization in lung, the effects of a mechanism-based inhibitor, 12-hydroxy-16-heptadecynoic acid (12-HHDYA) on PGE1 omega-hydroxylation, and the structure-function determinants which govern the activities of the enzymes encoded by this gene subfamily.