979 results on '"Arachidonate Lipoxygenases"'
Search Results
2. Biotransformation of C20- and C22-polyunsaturated fatty acids to 11S- and 13S-hydroxy fatty acids by Escherichia coli expressing 11S-lipoxygenase from Enhygromyxa salina
- Author
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Jin Lee, Tae-Hun Kim, Kyung-Chul Shin, Tae-Eui Lee, Min-Ju Kim, and Deok-Kun Oh
- Subjects
Fatty Acids ,Hydroxyeicosatetraenoic Acids ,Lipoxygenase ,Escherichia coli ,Fatty Acids, Unsaturated ,Bioengineering ,Cysteine ,Myxococcales ,General Medicine ,Arachidonate Lipoxygenases ,Applied Microbiology and Biotechnology ,Biotransformation ,Biotechnology - Abstract
Peroxidation and reduction of 11S- and 13S-positions on C20 and C22 polyunsaturated fatty acids (PUFAs) by Escherichia coli expressing highly active arachidonate (ARA) 11S-lipoxygenase (11S-LOX) from Enhygromyxa salina with the reducing agent cysteine.The specific activity and catalytic efficiency of ARA 11S-LOX from E. salina were 4.1- and 91-fold higher than those of only reported ARA 11S-LOX from Myxococcus xanthus, respectively. The hydroxy fatty acids (HFAs) obtained by the biotransformation of ARA, eicosapentaenoic acid (EPA), docosapentaenoic acid (DPA), and docosahexanoic acid (DHA) by Escherichia coli expressing 11S-LOX from E. salina in the presence of cysteine were identified as 11S-hydroxyeicosatetraenoic acid (11S-HETE), 11S-hydroxyeicosapentaenoic acid (11S-HEPE), 13S-hydroxydocosapentaenoic acid (13S-HDPA), and 13S-hydroxydocosahexaenoic acid (13S-HDHA), respectively. The recombinant cells converted 3 mM of ARA, EPA, DPA, and DHA into 2.9 mM of 11S-HETE, 2.4 mM 11S-HEPE, 1. 9 mM 13S-HDPA, and 2.2 mM 13S-HDHA in 60, 80, 120, and 120 min, corresponding to productivities of 72.5, 40.4, 18.5, and 22.4 μM minWe report the highest concentrations, conversion yields, and productivities of 11S- and 13S-hydroxy fatty acids from C20- and C22-PUFAs achieved via E. coli expressing highly active E. salina 11S-LOX. more...
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- 2022
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Catalog
3. Studies from Barts Cancer Institute Have Provided New Data on Cancer (Arachidonate 15-lipoxygenase-mediated production of Resolvin D5n-3 DPA abrogates pancreatic stellate cell-induced cancer cell invasion).
- Abstract
A recent report from the Barts Cancer Institute in London, United Kingdom, discusses the role of specialized pro-resolving mediators (SPMs) in cancer-stroma cross-talk and invasion. The study found that pancreatic stellate cells (PSCs) and cancer-associated fibroblasts (CAFs) can produce and respond to SPMs, which are known to modulate inflammation and its resolution. Specifically, the researchers discovered that the production of a specific SPM called Resolvin D5n-3 DPA, mediated by the enzyme arachidonate-15-lipoxygenase (ALOX15), may regulate cancer cell invasion. The findings suggest that targeting this pathway could potentially impact the chemoresistance and prognosis of pancreatic ductal adenocarcinoma (PDAC). [Extracted from the article] more...
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- 2023
4. Growth State-Dependent Expression of Arachidonate Lipoxygenases in the Human Endothelial Cell Line EA.hy926
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Mohammad G. Sabbir, Jeffrey T. Wigle, Carla G. Taylor, and Peter Zahradka
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endothelial cells ,lipoxygenase ,phenotype ,bioactive lipids ,Endothelial Cells ,Humans ,General Medicine ,Endothelium ,RNA, Messenger ,Arachidonate Lipoxygenases ,Lipids ,Cell Line - Abstract
Endothelial cells regulate vascular homeostasis through the secretion of various paracrine molecules, including bioactive lipids, but little is known regarding the enzymes responsible for generating these lipids under either physiological or pathophysiological conditions. Arachidonate lipoxygenase (ALOX) expression was therefore investigated in confluent and nonconfluent EA.h926 endothelial cells, which represent the normal quiescent and proliferative states, respectively. mRNAs for ALOX15, ALOX15B, and ALOXE3 were detected in EA.hy926 cells, with the highest levels present in confluent cells compared to nonconfluent cells. In contrast, ALOX5, ALOX12, and ALOX12B mRNAs were not detected. At the protein level, only ALOX15B and ALOXE3 were detected but only in confluent cells. ALOXE3 was also observed in confluent human umbilical artery endothelial cells (HUAEC), indicating that its expression, although previously unreported, may be a general feature of endothelial cells. Exposure to laminar flow further increased ALOXE3 levels in EA.hy926 cells and HUAECs. The evidence obtained in this study indicates that proliferative status and shear stress are both important factors that mediate endothelial ALOX gene expression. The presence of ALOX15B and ALOXE3 exclusively in quiescent human endothelial cells suggests their activity likely contributes to the maintenance of a healthy endothelium. more...
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- 2022
5. Gly188Arg substitution eliminates substrate inhibition in arachidonate 11R-lipoxygenase
- Author
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Priit Eek, Nigulas Samel, Maarja Lipp, Kaspar Põldemaa, and Ivar Järving
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0301 basic medicine ,Allosteric regulation ,Glycine ,Biophysics ,Arachidonic Acids ,Arginine ,Arachidonate Lipoxygenases ,Biochemistry ,Substrate Specificity ,03 medical and health sciences ,chemistry.chemical_compound ,Lipoxygenase ,0302 clinical medicine ,Fatty acid binding ,Animals ,Humans ,Amino Acid Sequence ,Enzyme Inhibitors ,Molecular Biology ,chemistry.chemical_classification ,biology ,Substrate (chemistry) ,Active site ,Cell Biology ,030104 developmental biology ,Enzyme ,Amino Acid Substitution ,chemistry ,030220 oncology & carcinogenesis ,Mutagenesis, Site-Directed ,biology.protein ,Arachidonic acid ,Uncompetitive inhibitor ,Sequence Alignment - Abstract
Lipoxygenases (LOXs) are dioxygenases that catalyze the oxygenation of polyunsaturated fatty acids to hydroperoxyl derivates. These products are precursors for different lipid mediators which are associated with pathogenesis of various diseases such as asthma, atherosclerosis and cancer. Several LOXs suffer from substrate inhibition, a potential regulatory mechanism, yet it is unclear what is the cause of this phenomenon. One such enzyme is the coral 11R-LOX which displays a significant decrease in turnover rate at arachidonic acid concentrations above 30 μM. In this report, site-directed mutagenesis and inhibition assays were employed to shed light on the mechanism of substrate inhibition in 11R-LOX. We found that introduction of a positive charge to the active site entrance with Gly188Arg substitution completely eliminates the slow-down at higher substrate concentrations. Inhibition of 11R-LOX by its catalysis product, 11(R)-hydroperoxyeicosatetraenoic acid, suggests an uncompetitive mechanism. We reason that substrate inhibition in 11R-LOX is due to additional fatty acid binding by the enzyme:substrate complex at an allosteric site situated in the very vicinity of the active site entrance. more...
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- 2019
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6. Production of 8S- and 10S-hydroxy polyunsaturated fatty acids by recombinant Escherichia coli cells expressing mouse arachidonate 8S-lipoxygenase
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Deok-Kun Oh, Min-Ju Seo, Dae Wook Kim, Woo-Ri Kang, and Kyung-Chul Shin
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0106 biological sciences ,0301 basic medicine ,Stereochemistry ,Bioengineering ,Arachidonate Lipoxygenases ,01 natural sciences ,Applied Microbiology and Biotechnology ,law.invention ,Hydroxylation ,Mice ,03 medical and health sciences ,chemistry.chemical_compound ,Lipoxygenase ,law ,010608 biotechnology ,Escherichia coli ,Animals ,chemistry.chemical_classification ,Recombinant escherichia coli ,Ethanol ,biology ,Temperature ,Substrate (chemistry) ,General Medicine ,Hydrogen-Ion Concentration ,Recombinant Proteins ,030104 developmental biology ,chemistry ,Fatty Acids, Unsaturated ,biology.protein ,Recombinant DNA ,Arachidonic acid ,Biotechnology ,Polyunsaturated fatty acid - Abstract
To quantitatively hydroxylate 8S- and 10S-positions on polyunsaturated fatty acids by recombinant Escherichia coli cells expressing mouse arachidonate 8S-lipoxygenase (8S-LOX). Hydroxylated products gained from the conversion of arachidonic acid (20:4Δ5Z,8Z,11Z,14Z, AA), eicosapentanoic acid (20:5Δ5Z,8Z,11Z,14Z,17Z, EPA), and (22:6Δ4Z,7Z,10Z,13Z,16Z,19Z, DHA) by recombinant E. coli cells containing 8S-LOX from mouse were identified as 8S-hydroxy-5,9,11,14(Z,E,Z,Z)-eicosatetranoic acid (8S-HETE), 8S-hydroxy-5,9,11,14,17(Z,E,Z,Z,Z)-eicosapentanoic acid (8S-HEPE), and 10S-hydroxy-4,8,12,14,16,19(Z,E,Z,Z,Z,Z)-docosahexaenoic acid (10S-HDoHE), respectively. Under the optimal hydroxylation conditions of pH 7.5, 30 °C, 5% (v/v) ethanol, 15 g cells l−1, and 5 mM substrate, AA, EPA, and DHA were hydroxylated into 4.37 mM 8S-HETE, 3.77 mM 8S-HEPE, and 3.13 mM 10S-HDoHE for 60, 90, and 60 min, with 87, 75, and 63% molar conversions, respectively. To the best of our knowledge, this is the first quantitatively biotechnological production of 8S-HETE, 8S-HEPE, and 10S-HDoHE. more...
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- 2019
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7. Regioselectivity of an arachidonate 9S-lipoxygenase from Sphingopyxis macrogoltabida that biosynthesizes 9S,15S- and 11S,17S-dihydroxy fatty acids from C20 and C22 polyunsaturated fatty acids
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Seong-Eun Kim, Jin Lee, Jung-Ung An, Tae-Hun Kim, Chae-Won Oh, Yoon-Joo Ko, Manigandan Krishnan, Joonhyeok Choi, Do-Young Yoon, Yangmee Kim, and Deok-Kun Oh
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Cell Biology ,Molecular Biology ,Arachidonate Lipoxygenases - Abstract
Lipoxygenases (LOXs) biosynthesize lipid mediators (LMs) as human signaling molecules. Among LMs, specialized pro-resolving mediators (SPMs) are involved in the resolution of inflammation and infection in humans. Here, the putative LOX from the bacterium Sphingopyxis macrogoltabida was identified as arachidonate 9S-LOX. The enzyme catalyzed oxygenation at the n-12 position of C20 and C22 polyunsaturated fatty acids (PUFAs) to form 9S- and 11S-hydroperoxy fatty acids, which were reduced to 9S- and 11S-hydroxy fatty acids (HFAs) by cysteine, respectively, and it catalyzed again oxygenation at the n-6 position of HFAs to form 9S,15S- and 11S,17S-DiHFAs, respectively. The regioselective residues of 9S-LOX were determined as lle395 and Val569 based on the amino acid alignment and homology models. The regioselectivity of the I395F variant was changed from the n-12 position on C20 PUFA to the n-6 position to form 15S-HFAs. This may be due to the reduction of the substrate-binding pocket by replacing the smaller Ile with a larger Phe. The V569W variant had a significantly lower second‑oxygenating activity compared to wild-type 9S-LOX because the insertion of the hydroxyl group of the first‑oxygenating products at the active site was seemed to be hindered by substituting a larger Trp for a smaller Val. The compounds, 11S-hydroxydocosapentaenoic acid, 9S,15S-dihydroxyeicosatetraenoic acid, 9S,15S-dihydroxyeicosapentaenoic acid, 11S,17S-hydroxydocosapentaenoic acid, and 11S,17S-dihydroxydocosahexaenoic acid, were newly identified by polarimeter, LC-MS/MS, and NMR. 11S,17S-DiHFAs as SPM isomers biosynthesized from C22 PUFAs showed anti-inflammatory activities in mouse and human cells. Our study contributes may stimulate physiological studies by providing new LMs. more...
- Published
- 2021
8. Characterization of Arachidonate 5S-Lipoxygenase from Danio rerio with High Activity for the Production of 5S- and 7S-Hydroxy Polyunsaturated Fatty Acids.
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Shin KC, Lee J, and Oh DK
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- Animals, Chromatography, Liquid, Tandem Mass Spectrometry, Fatty Acids, Unsaturated, Arachidonic Acid metabolism, Hydroxyeicosatetraenoic Acids metabolism, Docosahexaenoic Acids, Arachidonate 15-Lipoxygenase, Zebrafish metabolism, Arachidonate Lipoxygenases
- Abstract
A recombinant putative lipoxygenase (LOX) from Danio rerio (zebrafish), ALOX3c protein with 6-histidine tag, was purified using affinity chromatography, with a specific activity of 17.2 U mg
uperscript>-1 for arachidonic acid (AA). The molecular mass of the native ALOX3c was 156 kDa composed of a 78-kDa dimer by gel-filtration chromatography. The product obtained from the conversion of AA was identified as 5S-hydroxyeicosatetraenoic acid (5S-HETE) by HPLC and LC-MS/MS analyses. The specific activity and catalytic efficiency of the LOX from D. rerio for polyunsaturated fatty acids (PUFAs) followed the order AA (17.2 U mg-1 , 1.96 s-1 μM-1 ) > docosahexaenoic acid (DHA, 13.6 U mg-1 , 0.91 s-1 μM-1 ) > eicosapentaenoic acid (EPA, 10.5 U mg-1 , 0.65 s-1 μM-1 ) and these values for AA were the highest among the 5S-LOXs reported to date. Based on identified products and substrate specificity, the enzyme is an AA 5S-LOX. The enzyme exhibited the maximal activity at pH 8.0 and 20 °C with 0.1 mM Zn2+ in the presence of 10 mM cysteine. Under these reaction conditions, 6.88 U mL-1 D. rerio 5S-LOX converted 1.0 mM of AA, EPA, and DHA to 0.91 mM 5S-HETE, 0.72 mM 5S-hydroxyeicosapentaenoic acid (5S-HEPE), and 0.68 mM 7S-hydroxydocosahexaenoic acid (7S-HDHA) in 25, 30, and 25 min, corresponding to molar conversion rates of 91, 72, and 68% and productivities of 2.18, 1.44, and 1.63 mM h-1 , respectively. To the best of our knowledge, this study is the first to describe the bioconversion into 5S-HETE, 5S-HEPE, and 7S-HDHA for the application of biotechnological production., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.) more...- Published
- 2023
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9. Researchers at Korea University College of Medicine Zero in on Peripheral Artery Disease (Relationship between Arachidonate 5-Lipoxygenase-Activating Protein Gene and Peripheral Arterial Disease in Elderly Patients Undergoing General Surgery: A ...).
- Abstract
For more information on this research see: Relationship between Arachidonate 5-Lipoxygenase-Activating Protein Gene and Peripheral Arterial Disease in Elderly Patients Undergoing General Surgery: A Retrospective Observational Study. Keywords: Arachidonate 5-Lipoxygenase; Arachidonate Lipoxygenases; Cardiovascular Diseases and Conditions; Carrier Proteins; Dioxygenases; Enzymes and Coenzymes; Genetics; Health and Medicine; Iron-Binding Proteins; Lipoxygenase; Metalloproteins; Nonheme Iron Proteins; Oxidoreductases; Peripheral Artery Disease; Risk and Prevention; Surgery EN Arachidonate 5-Lipoxygenase Arachidonate Lipoxygenases Cardiovascular Diseases and Conditions Carrier Proteins Dioxygenases Enzymes and Coenzymes Genetics Health and Medicine Iron-Binding Proteins Lipoxygenase Metalloproteins Nonheme Iron Proteins Oxidoreductases Peripheral Artery Disease Risk and Prevention Surgery 2023 FEB 6 (NewsRx) -- By a News Reporter-Staff News Editor at Medical Devices & Surgical Technology Week -- Researchers detail new data in peripheral artery disease. [Extracted from the article] more...
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- 2023
10. Production of C20 9S- and C22 11S-hydroxy fatty acids by cells expressing Shewanella hanedai arachidonate 9S-lipoxygenase.
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Kim MJ, Lee J, Kim SE, Shin KC, and Oh DK
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- Fatty Acids, Unsaturated, Lipoxygenase, Arachidonate Lipoxygenases, Fatty Acids
- Abstract
The putative lipoxygenase (LOX) from the proteobacterium Shewanella hanedai was determined to be an 82 kDa monomeric enzyme by SDS-PAGE and gel filtration chromatography analysis. LOX was identified as a single-dioxygenating arachidonate (ARA) 9S-LOX by analyzing ARA-derived bioconversion products using high-performance liquid chromatography with reverse-, normal-, and chiral-phase columns and evaluating kinetic parameters for C20- and C22-polyunsaturated fatty acids (PUFAs). The catalytic efficiency (k
cat /Km ) values of 9S-LOX from S. hanedai for ARA, eicosapentaenoic acid, and docosahexaenoic acid were 3.1-, 4.1-, and 2.5-fold higher, respectively, than those only reported 9S-LOX from Sphingopyxis macrogoltabida with double-dioxygenating activity. To promote the production of C20 9S- and C22 11S-hydroxy fatty acids (HFAs) using Escherichia coli expressing 9S-LOX from S. hanedai, bioconversion conditions, including temperature, pH, solvent type and its concentration, concentrations of cells, and substrate, were optimized to 25 °C, pH 8.5, 6% (v/v) dimethyl sulfoxide, 0.2 g/l cells, and 7 mM ARA as substrate in a 500 ml-Erlenmeyer baffled flask with 50 ml reaction solution with agitation at 200 rpm in the presence of 10 mM cysteine as a reduction agent, respectively. Under these conditions, 6.4 mM 9S-hydroxyeicosatetraenoic acid, 6.2 mM 9S-hydroxyeicosapentaenoic acid, and 5.9 mM 11S-hydroxydocosahexaenoic acid were produced in 30 min, 40 min, and 60 min with specific productivities of 1067 μmol/min/g, 775 μmol/min/g, and 492 μmol/min/g, volumetric productivities of 213 μM/min, 155 μM/min, and 98 μM/min, and conversion yields of 91.4%, 88.6%, and 84.3%, respectively. To date, these are the highest specific productivities reported for the bioconversion of C20- and C22-PUFAs into HFAs. KEY POINTS: • Lipoxygenase from Shewanella hanedai was identified as arachidonate 9S-lipoxygenase • Optimization led to increased production of C20 9S- and C22 11S-hydroxy fatty acids • We reported the highest specific productivities of C20- and C22-hydroxy fatty acids., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.) more...- Published
- 2023
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11. Free docosahexaenoic acid promotes ferroptotic cell death via lipoxygenase dependent and independent pathways in cancer cells
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Kai Shan, Ninghan Feng, Doudou Zhu, Hongyan Qu, Guoling Fu, Jiaqi Li, Jing Cui, Heyan Chen, Rong Wang, Yumin Qi, and Yong Q. Chen
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Male ,Lipid Peroxides ,Nutrition and Dietetics ,Docosahexaenoic Acids ,Cell Death ,Lipoxygenase ,Medicine (miscellaneous) ,Lysophosphatidylcholines ,Lipoxygenases ,Arachidonate Lipoxygenases ,Carbon ,Cell Line, Tumor ,Neoplasms ,Humans ,Coenzyme A ,Lipid Peroxidation ,RNA, Small Interfering ,Reactive Oxygen Species ,Acyltransferases - Abstract
Ferroptosis is a form of regulated cell death that has the potential to be targeted as a cancer therapeutic strategy. But cancer cells have a wide range of sensitivities to ferroptosis, which limits its therapeutic potential. Accumulation of lipid peroxides determines the occurrence of ferroptosis. However, the type of lipid involved in peroxidation and the mechanism of lipid peroxide accumulation are less studied.The effects of fatty acids (10 μM) with different carbon chain length and unsaturation on ferroptosis were evaluated by MTT and LDH release assay in cell lines derived from prostate cancer (PC3, 22RV1, DU145 and LNCaP), colorectal cancer (HT-29), cervical cancer (HeLa) and liver cancer (HepG2). Inhibitors of apoptosis, necroptosis, autophagy and ferroptosis were used to determine the type of cell death. Then the regulation of reactive oxygen species (ROS) and lipid peroxidation by docosahexaenoic acid (DHA) was measured by HPLC-MS and flow cytometry. The avtive form of DHA was determined by siRNA mediated gene silencing. The role of lipoxygenases was checked by inhibitors and gene silencing. Finally, the effect of DHA on ferroptosis-mediated tumor killing was verified in xenografts.The sensitivity of ferroptosis was positively correlated with the unsaturation of exogenously added fatty acid. DHA (22:6 n-3) sensitized cancer cells to ferroptosis-inducing reagents (FINs) at the highest level in vitro and in vivo. In this process, DHA increased ROS accumulation, lipid peroxidation and protein oxidation independent of its membrane receptor, GPR120. Inhibition of long chain fatty acid-CoA ligases and lysophosphatidylcholine acyltransferases didn't affect the role of DHA. DHA-involved ferroptosis can be induced in both arachidonate lipoxygenase 5 (ALOX5) negative and positive cells. Down regulation of ALOX5 inhibited ferroptosis, while overexpression of ALOX5 promoted ferroptosis.DHA can effectively promote ferroptosis-mediated tumor killing by increasing intracellular lipid peroxidation. Both ALOX5 dependent and independent pathways are involved in DHA-FIN induced ferroptosis. And during this process, free DHA plays an important role. more...
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- 2020
12. Flipped regiospecificity in L434F mutant of 8-lipoxygenase
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Vipin Kumar Mishra and Sabyashachi Mishra
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0301 basic medicine ,Stereochemistry ,Protein Conformation ,Mutant ,General Physics and Astronomy ,chemistry.chemical_element ,Molecular Dynamics Simulation ,010402 general chemistry ,Crystallography, X-Ray ,01 natural sciences ,Oxygen ,Arachidonate Lipoxygenases ,03 medical and health sciences ,chemistry.chemical_compound ,Stereospecificity ,Molecule ,Phenyl group ,Animals ,Physical and Theoretical Chemistry ,chemistry.chemical_classification ,Quenching (fluorescence) ,Chemistry ,Substrate (chemistry) ,0104 chemical sciences ,030104 developmental biology ,Enzyme ,Mutation - Abstract
Lipoxygenases are non-heme iron containing enzymes that catalyze oxygenation of poly-unsaturated fatty acids in different animal and plant species with extremely high regio- and stereospecificity. Nature employs 8-lipoxygenase to produce 8R-hydroperoxide from the oxygenation of arachidonic acid. A single-point L434F mutation of 8-lipoxygenase alters the regio- and stereospecificity of the final products, with a product ratio of 66 : 34 for 8R- and 12S-hydroperoxide, respectively. A molecular level explanation of this flipped regiospecificity is presented in this work on the basis of molecular dynamics simulations and transition network analysis of oxygen migration in the protein matrix. Phe434 is shown to exist in two conformations, the so-called open and closed conformations. In the closed conformation, the phenyl group of Phe434 shields the C8 site of the substrate, thereby preventing access of the oxygen molecule to this site, which leads to a quenching of the 8R-product. On the other hand, both closed and open conformations of Phe434 allow the oxygen molecule to approach the pro-S face of the C12 site of the substrate, which enhances the propensity of the 12S-hydroperoxide. more...
- Published
- 2020
13. Human lipoxygenase isoforms form complex patterns of double and triple oxygenated compounds from eicosapentaenoic acid
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Stefan F. Kirsch, Martin Jübermann, Hartmut Kühn, Michael Rothe, Katharina M. Rund, Kateryna Goloshchapova, Wolf-Hagen Schunck, Nils Helge Schebb, Laura Kutzner, and Maximilian Blum
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0301 basic medicine ,Hydroxylation ,Arachidonate Lipoxygenases ,03 medical and health sciences ,chemistry.chemical_compound ,Lipoxygenase ,0302 clinical medicine ,Biosynthesis ,Humans ,Protein Isoforms ,Molecular Biology ,chemistry.chemical_classification ,biology ,ALOX15B ,Fatty acid ,Cell Biology ,Lipid signaling ,Eicosapentaenoic acid ,Recombinant Proteins ,Oxygen ,030104 developmental biology ,Enzyme ,chemistry ,Biochemistry ,Eicosapentaenoic Acid ,030220 oncology & carcinogenesis ,biology.protein ,Arachidonic acid - Abstract
Lipoxygenases (ALOX) are lipid peroxidizing enzymes that catalyze the biosynthesis of pro- and anti-inflammatory lipid mediators and have been implicated in (patho-)physiological processes. In humans, six functional ALOX isoforms exist and their arachidonic acid oxygenation products have been characterized. Products include leukotrienes and lipoxins which are involved in the regulation of inflammation and resolution. Oxygenation of n3-polyunsaturated fatty acids gives rise to specialized pro-resolving mediators, e.g. resolvins. However, the catalytic activity of different ALOX isoforms can lead to a multitude of potentially bioactive products. Here, we characterized the patterns of oxygenation products formed by human recombinant ALOX5, ALOX15, ALOX15B and ALOX12 from eicosapentaenoic acid (EPA) and its 18-hydroxy derivative 18-HEPE with particular emphasis on double and triple oxygenation products. ALOX15 and ALOX5 formed a complex mixture of various double oxygenation products from EPA, which include 5,15-diHEPE and various 8,15-diHEPE isomers. Their biosynthetic mechanisms were explored using heavy oxygen isotopes (H218O, 18O2 gas) and three catalytic activities contributed to product formation: i) fatty acid oxygenase activity, ii) leukotriene synthase activity, iii) lipohydroperoxidase activity. For ALOX15B and ALOX12 more specific product patterns were identified, which was also the case when these enzymes reacted in concert with ALOX5. Several double oxygenated compounds were formed from 18-HEPE by ALOX5, ALOX15B and ALOX12 including previously identified resolvins (RvE2, RvE3), while formation of triple oxygenation products, e.g. 5,17,18-triHEPE, required ALOX5. Taken together our data show that EPA can be converted by human ALOX isoforms to a large number of secondary oxygenation products, which might exhibit bioactivity. more...
- Published
- 2020
14. Role of arachidonic acid lipoxygenase pathway in Asthma
- Author
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Yacan, Luo, Minli, Jin, Lejing, Lou, Song, Yang, Chengye, Li, Xi, Li, Meixi, Zhou, and Chang, Cai
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Lipoxins ,Pharmacology ,Leukotrienes ,Arachidonate 5-Lipoxygenase ,Physiology ,Humans ,Cell Biology ,Lipoxygenases ,Arachidonate Lipoxygenases ,Biochemistry ,Asthma - Abstract
The arachidonic acid (AA) metabolism pathways play a key role in immunological response and inflammation diseases, such as asthma, etc. AA in cell membranes can be metabolized by lipoxygenases (LOXs) to a screen of bioactive substances that include leukotrienes (LTs), lipoxins (LXs), and eicosatetraenoic acids (ETEs), which are considered closely related to the pathophysiology of respiratory allergic disease. Studies also verified that drugs regulating AA LOXs pathway have better rehabilitative intervention for asthma. This review aims to summarize the physiological and pathophysiological importance of AA LOXs metabolism pathways in asthma and to discuss its prospects of therapeutic strategies. more...
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- 2022
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15. Biotransformation of polyunsaturated fatty acids to bioactive hepoxilins and trioxilins by microbial enzymes
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Do-Young Yoon, Yoon-Joo Ko, Deok-Kun Oh, Jung-Ung An, Yong-Seok Song, and Kyoung-Rok Kim
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0301 basic medicine ,Myxococcus xanthus ,Science ,General Physics and Astronomy ,Arachidonate 12-Lipoxygenase ,Arachidonate Lipoxygenases ,General Biochemistry, Genetics and Molecular Biology ,Article ,03 medical and health sciences ,chemistry.chemical_compound ,0302 clinical medicine ,8,11,14-Eicosatrienoic Acid ,Biotransformation ,Bacterial Proteins ,Epoxide hydrolase ,lcsh:Science ,chemistry.chemical_classification ,Epoxide Hydrolases ,Multidisciplinary ,biology ,Molecular Structure ,Chemistry ,General Chemistry ,Peroxisome ,biology.organism_classification ,Eicosapentaenoic acid ,030104 developmental biology ,Biochemistry ,Docosahexaenoic acid ,Fatty Acids, Unsaturated ,Arachidonic acid ,lcsh:Q ,030217 neurology & neurosurgery ,Metabolic Networks and Pathways ,Polyunsaturated fatty acid - Abstract
Hepoxilins (HXs) and trioxilins (TrXs) are involved in physiological processes such as inflammation, insulin secretion and pain perception in human. They are metabolites of polyunsaturated fatty acids (PUFAs), including arachidonic acid, eicosapentaenoic acid and docosahexaenoic acid, formed by 12-lipoxygenase (LOX) and epoxide hydrolase (EH) expressed by mammalian cells. Here, we identify ten types of HXs and TrXs, produced by the prokaryote Myxococcus xanthus, of which six types are new, namely, HXB5, HXD3, HXE3, TrXB5, TrXD3 and TrXE3. We succeed in the biotransformation of PUFAs into eight types of HXs (>35% conversion) and TrXs (>10% conversion) by expressing M. xanthus 12-LOX or 11-LOX with or without EH in Escherichia coli. We determine 11-hydroxy-eicosatetraenoic acid, HXB3, HXB4, HXD3, TrXB3 and TrXD3 as potential peroxisome proliferator-activated receptor-γ partial agonists. These findings may facilitate physiological studies and drug development based on lipid mediators., Hepoxilins (HXs) and trioxilins (TrXs) are lipid metabolites with roles in inflammation and insulin secretion. Here, the authors discover a prokaryotic source of HXs and TrXs, identify the biosynthetic enzymes and heterologously express HXs and TrXs in E. coli. more...
- Published
- 2018
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16. Moisturizers change the mRNA expression of enzymes synthesizing skin barrier lipids.
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Buraczewska, Izabela, Berne, Berit, Lindberg, Magnus, Lodén, Marie, and Törmä, Hans
- Subjects
- *
MESSENGER RNA , *ISOPENTENOIDS , *GENE expression , *SKIN diseases , *DERMATOLOGY - Abstract
In a previous study, 7-week treatment of normal human skin with two test moisturizers, Complex cream and Hydrocarbon cream, was shown to affect mRNA expression of certain genes involved in keratinocyte differentiation. Moreover, the treatment altered transepidermal water loss (TEWL) in opposite directions. In the present study, the mRNA expression of genes important for formation of barrier lipids, i.e., cholesterol, free fatty acids and ceramides, was examined. Treatment with Hydrocarbon cream, which increased TEWL, also elevated the gene expression of GBA, SPTLC2, SMPD1, ALOX12B, ALOXE3, and HMGCS1. In addition, the expression of PPARG was decreased. On the other hand, Complex cream, which decreased TEWL, induced only the expression of PPARG, although not confirmed at the protein level. Furthermore, in the untreated skin, a correlation between the mRNA expression of PPARG and ACACB, and TEWL was found, suggesting that these genes are important for the skin barrier homeostasis. The observed changes further demonstrate that long-term treatment with certain moisturizers may induce dysfunctional skin barrier, and as a consequence several signaling pathways are altered. [ABSTRACT FROM AUTHOR] more...
- Published
- 2009
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17. Origin of Regio- and Stereospecific Catalysis by 8-Lipoxygenase
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Sabyashachi Mishra and Vipin Kumar Mishra
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Allylic rearrangement ,Stereochemistry ,Protein Conformation ,chemistry.chemical_element ,Molecular Dynamics Simulation ,010402 general chemistry ,Hydrogen atom abstraction ,01 natural sciences ,Oxygen ,Arachidonate Lipoxygenases ,Catalysis ,Substrate Specificity ,Stereospecificity ,0103 physical sciences ,Materials Chemistry ,Physical and Theoretical Chemistry ,chemistry.chemical_classification ,010304 chemical physics ,Substrate (chemistry) ,Stereoisomerism ,0104 chemical sciences ,Surfaces, Coatings and Films ,Enzyme ,chemistry ,Mutation ,Biocatalysis ,Carbon - Abstract
Lipoxygenases (lox's) are a group of non-heme iron containing enzymes that catalyze oxygenation of polyunsaturated fatty acids with precise regio- and stereoselectivities. The origin of regio- and stereospecific catalysis by 8-lox is explored in its wild-type (wt) form and in three mutants (Arg185Ala, Ala592Met, and Ala623His). The catalytic action of this enzyme progresses in two steps, namely, hydrogen abstraction from one double allylic carbon atom of substrate followed by oxygen insertion at the resulting prochiral carbon radical of the substrate. It is shown that the positional specificity of the hydrogen abstraction is a result of conformational dynamics of the bound substrate. While the C10 atom of the substrate is found to be the most probable site of hydrogen abstraction in the wt-lox, hydrogen abstraction from C13 is more favorable in the mutants. The present study discovers the presence of an interconnected network of a three-channel migration pathway operating in the protein matrix for efficient oxygen transport. Each migration channel is bestowed with a pocket at the peripheral region of protein as an oxygen access site, which transfers the oxygen to the active site through a well-connected migration path on a time scale of a few hundred picoseconds. By a careful geometric analysis of the oxygen pockets near the substrate binding cleft, the present study identifies the launching sites for oxygenation at the prochiral carbon centers C8, C11, C12, and C15 and the stereochemistry ( more...
- Published
- 2019
18. Fatty acids modulate the expression of pyruvate kinase and arachidonate-lipoxygenase through PPARγ/CYP2C45 pathway: a link to goose fatty liver
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Tuoyu Geng, Daoqing Gong, Qinghang Wang, R Zhang, Tongjun Liu, M.M. Zhao, and L Liu
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Male ,Pyruvate Kinase ,Peroxisome proliferator-activated receptor ,Arachidonate Lipoxygenases ,Avian Proteins ,03 medical and health sciences ,Goose ,Cytochrome P-450 Enzyme System ,biology.animal ,Geese ,medicine ,Animals ,Glycolysis ,Poultry Diseases ,030304 developmental biology ,chemistry.chemical_classification ,0303 health sciences ,biology ,Base Sequence ,Fatty liver ,Fatty Acids ,0402 animal and dairy science ,Troglitazone ,Fatty acid ,04 agricultural and veterinary sciences ,General Medicine ,medicine.disease ,040201 dairy & animal science ,Fatty Liver ,PPAR gamma ,Biochemistry ,chemistry ,Arachidonate 5-lipoxygenase ,biology.protein ,Animal Science and Zoology ,Pyruvate kinase ,medicine.drug ,Signal Transduction - Abstract
Cytochrome P-450 2C45 (CYP2C45) is the most highly expressed cytochrome P-450 isoform in chicken liver, and may play an important role in avian liver biology. However, information regarding the function of CYP2C45 in fatty liver is generally limited. The aim of this study was to investigate the role of CYP2C45 during the development of goose fatty liver. Our result indicated that the transcription of CYP2C45, together with PK and ALOX5, was increased in goose liver upon overfeeding for 19 D (P < 0.05). In goose primary hepatocytes, CYP2C45 RNA expression was also upgraded by the treatment with various chemicals like insulin, the fatty acids, and PPAR agonists (P < 0.05). We also found that both CYP2C45 overexpression and troglitazone treatment could increase the expression of pyruvate kinase (PK) and arachidonate 5-lipoxygenase (ALOX5), and furthermore, showed that the up-regulation of PK and ALOX5 induced by troglitazone could be suppressed by small interfering RNAs targeting CYP2C45 (P < 0.05). These findings suggest that fatty acids treatment and the overfeeding can induce the up-regulation of CYP2C45 expression possibly via PPARγ and that the induction of PK and ALOX5 in goose fatty liver is at least partially attributed to fatty acid-induced expression of CYP2C45. Thus, our data provides an insight into the mechanism by which glycolysis and arachidonic acid metabolism are modulated in goose fatty liver. more...
- Published
- 2019
19. Role of arachidonic acid lipoxygenase pathway in Asthma.
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Luo Y, Jin M, Lou L, Yang S, Li C, Li X, Zhou M, and Cai C
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- Arachidonate 5-Lipoxygenase, Arachidonate Lipoxygenases, Humans, Leukotrienes, Lipoxygenases, Asthma drug therapy, Lipoxins
- Abstract
The arachidonic acid (AA) metabolism pathways play a key role in immunological response and inflammation diseases, such as asthma, etc. AA in cell membranes can be metabolized by lipoxygenases (LOXs) to a screen of bioactive substances that include leukotrienes (LTs), lipoxins (LXs), and eicosatetraenoic acids (ETEs), which are considered closely related to the pathophysiology of respiratory allergic disease. Studies also verified that drugs regulating AA LOXs pathway have better rehabilitative intervention for asthma. This review aims to summarize the physiological and pathophysiological importance of AA LOXs metabolism pathways in asthma and to discuss its prospects of therapeutic strategies., (Copyright © 2021 Elsevier Inc. All rights reserved.) more...
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- 2022
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20. Expression of an 8 R -Lipoxygenase From the Coral Plexaura homomalla
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Nathaniel C. Gilbert, David B. Neau, and Marcia E. Newcomer
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0301 basic medicine ,Coral ,Crystallography, X-Ray ,Arachidonate Lipoxygenases ,Article ,Substrate Specificity ,03 medical and health sciences ,Lipoxygenase ,Protein Domains ,Escherichia coli ,Animals ,Enzyme Assays ,chemistry.chemical_classification ,Enzyme substrate complex ,Arachidonic Acid ,Binding Sites ,030102 biochemistry & molecular biology ,biology ,Plexaura homomalla ,fungi ,Substrate (chemistry) ,Anthozoa ,biology.organism_classification ,Recombinant Proteins ,030104 developmental biology ,Enzyme ,Metabolic Engineering ,chemistry ,Biochemistry ,biology.protein - Abstract
Methods are presented for the use of the coral 8R-lipoxygenase from the Caribbean sea whip coral Plexaura homomalla as a model enzyme for structural studies of animal lipoxygenases. The 8R-lipoxygenase is remarkably stable and can be stored at 4°C for 3 months with virtually no loss of activity. In addition, an engineered “pseudo wild-type” enzyme is soluble in the absence of detergents, which helps facilitate the preparation of enzyme:substrate complexes. more...
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- 2018
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21. Gaining insight into the chemistry of lipoxygenases: a computational investigation into the catalytic mechanism of (8R)-lipoxygenase
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James W. Gauld, Riam Jamil, and Eric A. C. Bushnell
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Peroxidation ,Protein Conformation ,Stereochemistry ,Lipoxygenase ,Quantum mechanical/molecular mechanical ,Arachidonate Lipoxygenases ,Biochemistry ,Inorganic Chemistry ,Active center ,Electron transfer ,General chemistry ,Catalytic Domain ,Animals ,Reactivity (chemistry) ,Biochemistry, Biophysics, and Structural Biology ,Non-heme iron enzyme ,biology ,Chemistry ,Mechanism (biology) ,Active site ,Anthozoa ,Peroxides ,Multistate reactivity ,Enzyme Activation ,Molecular Docking Simulation ,biology.protein ,Quantum Theory ,Soybeans ,(8R)-Lipoxygenases ,Function (biology) - Abstract
Lipoxygenases (LOXs) are ubiquitous in nature and catalyze a range of life-essential reactions within organisms. In particular they are critical to the formation of eicosanoids, which are critical for normal cell function. However, a number of important questions about the reactivity and mechanism of these enzymes still remain. Specifically, although the initial step in the mechanism of LOXs has been well studied, little is known of subsequent steps. Thus, with use of a quantum mechanical/molecular mechanical approach, the complete catalytic mechanism of (8R)-LOX was investigated. The results have provided a better understanding of the general chemistry of LOXs as a whole. In particular, from comparisons with soybean LOX-1, it appears that the initial proton-coupled electron transfer may be very similar among all LOXs. Furthermore, LOXs appear to undergo multistate reactivity where potential spin inversion of an electron may occur either in the attack of O2 or in the regeneration of the active site. Lastly, it is shown that with the explicit modeling of the environment, the regeneration of the active center likely occurs via the rotation of the intermediate followed by an outer-sphere H⋅" role="presentation" style="box-sizing: border-box; display: inline-table; line-height: normal; letter-spacing: normal; word-spacing: normal; overflow-wrap: normal; white-space: nowrap; float: none; direction: ltr; max-width: none; max-height: none; min-width: 0px; min-height: 0px; border: 0px; padding: 0px; margin: 0px; position: relative;">H⋅H⋅ transfer as opposed to the formation of a “purple intermediate” complex. more...
- Published
- 2013
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22. Effect of N-Feruloylserotonin and Methotrexate on Severity of Experimental Arthritis and on Messenger RNA Expression of Key Proinflammatory Markers in Liver
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Pauliková I, Danica Mihalova, Iveta Hrádková, Viera Kuncirova, Ľudmila Pašková, Radomír Nosáľ, Tomáš Čavojský, Juraj Harmatha, Lýdia Bezáková, František Bilka, Katarína Šišková, Silvester Ponist, and Katarína Bauerová more...
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0301 basic medicine ,Male ,Time Factors ,Arthritis ,Arachidonate Lipoxygenases ,Severity of Illness Index ,Immunology and Allergy ,biology ,General Medicine ,medicine.anatomical_structure ,C-Reactive Protein ,Liver ,Organ Specificity ,Rheumatoid arthritis ,Cytokines ,medicine.symptom ,Inflammation Mediators ,medicine.drug ,Research Article ,lcsh:Immunologic diseases. Allergy ,musculoskeletal diseases ,medicine.medical_specialty ,Serotonin ,Combination therapy ,Article Subject ,Immunology ,Spleen ,Inflammation ,Proinflammatory cytokine ,03 medical and health sciences ,Internal medicine ,medicine ,Animals ,business.industry ,C-reactive protein ,medicine.disease ,Arthritis, Experimental ,Rats ,Disease Models, Animal ,030104 developmental biology ,Endocrinology ,Methotrexate ,Gene Expression Regulation ,biology.protein ,business ,lcsh:RC581-607 ,Transcriptome ,Biomarkers - Abstract
Rheumatoid arthritis (RA) is a chronic inflammatory disease, leading to progressive destruction of joints and extra-articular tissues, including organs such as liver and spleen. The purpose of this study was to compare the effects of a potential immunomodulator, natural polyphenol N-feruloylserotonin (N-f-5HT), with methotrexate (MTX), the standard in RA therapy, in the chronic phase of adjuvant-induced arthritis (AA) in male Lewis rats. The experiment included healthy controls (CO), arthritic animals (AA), AA given N-f-5HT (AA-N-f-5HT), and AA given MTX (AA-MTX). N-f-5HT did not affect the body weight change and clinical parameters until the 14th experimental day. Its positive effect was rising during the 28-day experiment, indicating a delayed onset of N-f-5HT action. Administration of either N-f-5HT or MTX caused reduction of inflammation measured as the level of CRP in plasma and the activity of LOX in the liver. mRNA transcription of TNF-αand iNOS in the liver was significantly attenuated in both MTX and N-f-5HT treated groups of arthritic rats. Interestingly, in contrast to MTX, N-f-5HT significantly lowered the level of IL-1βin plasma and IL-1βmRNA expression in the liver and spleen of arthritic rats. This speaks for future investigations of N-f-5HT as an agent in the treatment of RA in combination therapy with MTX. more...
- Published
- 2016
23. Structure of a Calcium-dependent 11R-Lipoxygenase Suggests a Mechanism for Ca2+ Regulation
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Marcia E. Newcomer, Priit Eek, Reet Järving, Nigulas Samel, Nathaniel C. Gilbert, and Ivar Järving
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Allosteric regulation ,Molecular Conformation ,chemistry.chemical_element ,Calcium ,Crystallography, X-Ray ,Arachidonate Lipoxygenases ,Biochemistry ,Mass Spectrometry ,Lipoxygenase ,Protein structure ,Catalytic Domain ,Calcium-binding protein ,Animals ,Humans ,Molecular Biology ,Chromatography, High Pressure Liquid ,biology ,Calcium-Binding Proteins ,Cell Membrane ,Peripheral membrane protein ,Active site ,Cell Biology ,Anthozoa ,Protein Structure, Tertiary ,Kinetics ,Gene Expression Regulation ,Models, Chemical ,chemistry ,Protein Structure and Folding ,Liposomes ,Mutagenesis, Site-Directed ,biology.protein ,Biophysics ,Eicosanoids ,Signal transduction ,Dimerization ,Allosteric Site - Abstract
Lipoxygenases (LOXs) are a key part of several signaling pathways that lead to inflammation and cancer. Yet, the mechanisms of substrate binding and allosteric regulation by the various LOX isoforms remain speculative. Here we report the 2.47-Å resolution crystal structure of the arachidonate 11R-LOX from Gersemia fruticosa, which sheds new light on the mechanism of LOX catalysis. Our crystallographic and mutational studies suggest that the aliphatic tail of the fatty acid is bound in a hydrophobic pocket with two potential entrances. We speculate that LOXs share a common T-shaped substrate channel architecture that gives rise to the varying positional specificities. A general allosteric mechanism is proposed for transmitting the activity-inducing effect of calcium binding from the membrane-targeting PLAT (polycystin-1/lipoxygenase/α-toxin) domain to the active site via a conserved π-cation bridge. more...
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- 2012
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24. Cyclooxygenases and lipoxygenases in cancer
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Claus Schneider and Ambra Pozzi
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Cancer Research ,Chemokine ,Antineoplastic Agents ,Arachidonate Lipoxygenases ,Article ,Neoplasms ,medicine ,Animals ,Humans ,Cyclooxygenase Inhibitors ,Receptor ,biology ,Cell growth ,Cancer ,medicine.disease ,Extravasation ,Cell Transformation, Neoplastic ,Linoleic Acids ,Oncology ,Nuclear receptor ,Biochemistry ,Eicosanoid ,Prostaglandin-Endoperoxide Synthases ,biology.protein ,Cancer research ,Eicosanoids ,Cyclooxygenase - Abstract
Cancer initiation and progression are multistep events that require cell proliferation, migration, extravasation to the blood or lymphatic vessels, arrest to the metastatic site, and ultimately secondary growth. Tumor cell functions at both primary or secondary sites are controlled by many different factors, including growth factors and their receptors, chemokines, nuclear receptors, cell-cell interactions, cell-matrix interactions, as well as oxygenated metabolites of arachidonic acid. The observation that cyclooxygenases and lipoxygenases and their arachidonic acid-derived eicosanoid products (prostanoids and HETEs) are expressed and produced by tumor cells, together with the finding that these enzymes can regulate cell growth, survival, migration, and invasion, has prompted investigators to analyze the roles of these enzymes in cancer progression. In this review, we focus on the contribution of cyclooxygenase- and lipoxygenase-derived eicosanoids to tumor cell function in vitro and in vivo and discuss hope and tribulations of targeting these enzymes for cancer prevention and treatment. more...
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- 2011
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25. Role of Lipoxygenase Metabolites of Arachidonic Acid in Enhanced Pulmonary Artery Contractions of Female Rabbits
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Sandra L. Pfister
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Male ,medicine.medical_specialty ,Elevated pulmonary artery pressure ,Endothelium ,medicine.drug_class ,Blotting, Western ,Pulmonary Artery ,Biology ,Arachidonate Lipoxygenases ,Mass Spectrometry ,Article ,chemistry.chemical_compound ,Sex Factors ,medicine.artery ,Internal medicine ,Hydroxyeicosatetraenoic Acids ,Internal Medicine ,medicine ,Animals ,Analysis of Variance ,Arachidonic Acid ,medicine.disease ,Pulmonary hypertension ,Endocrinology ,medicine.anatomical_structure ,chemistry ,Vasoconstriction ,Estrogen ,Pulmonary artery ,Vascular resistance ,Female ,Arachidonic acid ,Endothelium, Vascular ,Rabbits ,medicine.symptom - Abstract
Pulmonary arterial hypertension is characterized by elevated pulmonary artery pressure and vascular resistance. In women the incidence is 4 fold greater than that in men. Studies suggest sustained vasoconstriction is a factor in increased vascular resistance. Possible vasoconstrictor mediators include arachidonic acid-derived lipoxygenase metabolites. Our studies in rabbits showed enhanced endothelium-dependent contractions to arachidonic acid in pulmonary arteries from females compared to males. Because treatment with a non-specific lipoxygenase inhibitor reduced contractions in females but not males, the present study identified which lipoxygenase isoform contributes to sex-specific pulmonary artery vasoconstriction. 15- and 5- but not 12-lipoxygenase protein expression was greater in females. Basal and A23187-stimulated release of 15-, 5- and 12-hydroxyeicosatetraenoic acid from females and males was measured by liquid chromatography/mass spectrometry. Only 15-hydroxyeicosatetraenoic acid synthesis was greater in females compared to males under both basal and stimulated conditions. Vascular contractions to 15-hydroxyeicosatetraenoic acid were enhanced in females compared to males (maximal contraction; 44 ± 6% vs 25 ± 3%). The specific 15-lipoxygenase inhibitor PD146176 (12 μmol/L) decreased arachidonic acid-induced contractions in females (maximal contraction; 93 ± 4% vs 57 ± 10%). If male pulmonary arteries were incubated with estrogen (1 μmol/L, 18 hrs), protein expression of 15-lipoxygenase, and 15-hydroxyeicosatetraenoic acid production increased. Mechanisms to explain the increased incidence of pulmonary hypertension in women are not known. Results suggest the 15-lipoxygenase pathway is different between females and males and is regulated by estrogen. Understanding this novel sex-specific mechanism may provide insight into the increased incidence of pulmonary hypertension in females. more...
- Published
- 2011
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26. Variation in eicosanoid genes, non-fatal myocardial infarction and ischemic stroke
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Kenneth Rice, Thomas Lumley, Kerri L. Wiggins, Joshua C. Bis, Kristin D. Marciante, Rozenn N. Lemaitre, Susan R. Heckbert, Bruce M. Psaty, and Nicholas L. Smith
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Adult ,Male ,medicine.medical_specialty ,Myocardial Infarction ,PTGS1 ,Prostaglandin E synthase ,Arachidonate Lipoxygenases ,Polymorphism, Single Nucleotide ,Risk Assessment ,Article ,Brain Ischemia ,Prostacyclin synthase ,Cytochrome P-450 Enzyme System ,Gene Frequency ,Risk Factors ,Internal medicine ,Odds Ratio ,medicine ,Humans ,Genetic Predisposition to Disease ,Myocardial infarction ,Stroke ,Aged ,Prostaglandin-E Synthases ,Aspirin ,biology ,business.industry ,Middle Aged ,medicine.disease ,Intramolecular Oxidoreductases ,Endocrinology ,Cyclooxygenase 2 ,Case-Control Studies ,Cyclooxygenase 1 ,Cardiology ,biology.protein ,Eicosanoids ,Platelet aggregation inhibitor ,Female ,Thromboxane-A Synthase ,Thromboxane-A synthase ,Cardiology and Cardiovascular Medicine ,business ,Platelet Aggregation Inhibitors ,medicine.drug - Abstract
Objectives Eicosanoids are lipid mediators that may play a role in atherosclerosis. We investigated the association of common genetic variation in prostaglandin H synthase 1 ( PTGS1 ), prostaglandin H synthase 2 ( PTGS2 ), thromboxane A2 synthase ( TBXAS1 ), prostacyclin synthase ( PTGIS ), prostaglandin E synthase ( PTGES ), 5-lipoxygenase activating protein ( ALOX5AP ), 12-lipoxygenase ( ALOX12 ) and 15-lipoxygenase ( ALOX15 ) with the risks of myocardial infarction (MI) and ischemic stroke. A secondary aim was to replicate the interaction of PTGS2 rs20417 (-765G to C) with aspirin use on coronary heart disease risk observed in the Atherosclerosis Risk in Communities Study (ARIC). Methods We conducted a case-control study in a large Health Maintenance Organization. Cases were men and women, aged 30–79 years with incident non-fatal myocardial infarction ( n =1063) or ischemic stroke ( n =469) between January 1995 and December 2004. Controls ( n =3462) were randomly selected and frequency matched to cases on age, sex, hypertension and calendar year. Results Common variation in TBXAS1 and PTGIS was associated with MI risk ( p -value for global Chi-square test, 0.01 and 0.03, respectively). Common variation in ALOX5AP , ALOX12 , ALOX15 , PTGS1 , PTGS2 and PTGES was not associated with risks of MI and ischemic stroke. We replicated the observation of the Atherosclerosis Risk in Communities Study and observed an interaction of rs20417 with aspirin use on myocardial infarction risk ( p for interaction=0.03). Conclusions Study results suggest that variation in TBXAS1 and PTGIS may influence MI risk, and carriers of rs20417C allele might derive greater benefits from aspirin use in primary prevention. more...
- Published
- 2009
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27. Cytochrome P450 and Lipoxygenase Metabolites on Renal Function
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John D. Imig and Md. Abdul Hye Khan
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medicine.medical_specialty ,Apoptosis ,urologic and male genital diseases ,Kidney ,Arachidonate Lipoxygenases ,Article ,chemistry.chemical_compound ,Cytochrome P-450 Enzyme System ,Internal medicine ,medicine ,Animals ,Humans ,biology ,Hemodynamics ,Kidney metabolism ,Cytochrome P450 ,Renal Reabsorption ,medicine.anatomical_structure ,Endocrinology ,chemistry ,Renal physiology ,Renal blood flow ,biology.protein ,Arachidonic acid ,Cyclooxygenase - Abstract
Arachidonic acid metabolites have a myriad of biological actions including effects on the kidney to alter renal hemodynamics and tubular transport processes. Cyclooxygenase metabolites are products of an arachidonic acid enzymatic pathway that has been extensively studied in regards to renal function. Two lesser-known enzymatic pathways of arachidonic acid metabolism are the lipoxygenase (LO) and cytochrome P450 (CYP) pathways. The importance of LO and CYP metabolites to renal hemodynamics and tubular transport processes is now being recognized. LO and CYP metabolites have actions to alter renal blood flow and glomerular filtration rate. Proximal and distal tubular sodium transport and fluid and electrolyte homeostasis are also significantly influenced by renal CYP and LO levels. Metabolites of the LO and CYP pathways also have renal actions that influence renal inflammation, proliferation, and apoptotic processes at vascular and epithelial cells. These renal LO and CYP pathway actions occur through generation of specific metabolites and cell-signaling mechanisms. Even though the renal physiological importance and actions for LO and CYP metabolites are readily apparent, major gaps remain in our understanding of these lipid mediators to renal function. Future studies will be needed to fill these major gaps regarding LO and CYP metabolites on renal function. more...
- Published
- 2016
28. Epidermal Lipoxygenase Products of the Hepoxilin Pathway Selectively Activate the Nuclear Receptor PPARα
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Claus Schneider, William E. Boeglin, Zheyong Yu, and Alan R. Brash
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Lipoxygenase ,Receptors, Cytoplasmic and Nuclear ,Peroxisome proliferator-activated receptor ,Arachidonate Lipoxygenases ,Biochemistry ,ALOXE3 ,Cell Line ,chemistry.chemical_compound ,8,11,14-Eicosatrienoic Acid ,Genes, Reporter ,Hydroxyeicosatetraenoic Acids ,Humans ,PPAR alpha ,12-Hydroxy-5,8,10,14-eicosatetraenoic Acid ,PPAR delta ,Luciferases ,Receptor ,chemistry.chemical_classification ,integumentary system ,biology ,Organic Chemistry ,Cell Differentiation ,Cell Biology ,Ligand (biochemistry) ,PPAR gamma ,Epidermal Cells ,Nuclear receptor ,chemistry ,Hepoxilin ,biology.protein ,Arachidonic acid ,Epidermis ,Signal Transduction - Abstract
Arachidonic acid can be transformed into a specific epoxyalcohol product via the sequential action of two epidermal lipoxygenases, 12R-LOX and eLOX3. Functional impairment of either lipoxygenase gene (ALOX12B or ALOXE3) results in ichthyosis, suggesting a role for the common epoxyalcohol product or its metabolites in the differentiation of normal human skin. Here we tested the ability of products derived from the epidermal LOX pathway to activate the peroxisome proliferator-activated receptors PPARalpha, gamma, and delta, which have been implicated in epidermal differentiation. Using a dual luciferase reporter assay in PC3 cells, the 12R-LOX/eLOX3-derived epoxyalcohol, 8R-hydroxy-11R,12R-epoxyeicosa-5Z,9E,14Z-trienoic acid, activated PPARalpha with similar in potency to the known natural ligand, 8S-hydroxyeicosatetraenoic acid (8S-HETE) (both at 10 microM concentration). In contrast, the PPARgamma and PPARdelta receptor isoforms were not activated by the epoxyalcohol. Activation of PPARalpha was also observed using the trihydroxy hydrolysis products (trioxilins) of the unstable epoxyalcohol. Of the four trioxilins isolated and characterized, the highest activation was observed with the isomer that is also formed by enzymatic hydrolysis of the epoxyalcohol. Formation of a ligand for the nuclear receptor PPARalpha may be one possibility by which 12R-LOX and eLOX3 contribute to epidermal differentiation. more...
- Published
- 2007
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29. Inducible expression of 15-lipoxygenase-2 and 8-lipoxygenase inhibits cell growth via common signaling pathways
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Peter Krieg, Gerhard Fürstenberger, and Dorothea Schweiger
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Keratinocytes ,Time Factors ,antioxidant ,p38 mitogen-activated protein kinases ,keratinocyte ,QD415-436 ,Biology ,Arachidonate Lipoxygenases ,p38 Mitogen-Activated Protein Kinases ,Biochemistry ,Gene Expression Regulation, Enzymologic ,Cell Line ,Linoleic Acid ,Mice ,chemistry.chemical_compound ,Endocrinology ,Hydroxyeicosatetraenoic Acids ,Animals ,Arachidonate 15-Lipoxygenase ,Humans ,Lipoxygenase Inhibitors ,Protein kinase A ,Cell Proliferation ,reactive oxygen species ,Arachidonic Acid ,Dose-Response Relationship, Drug ,Cell growth ,Kinase ,DNA ,Cell Biology ,Molecular biology ,Acetylcysteine ,Cell biology ,chemistry ,Cell culture ,Doxycycline ,mitogen-activated protein kinases ,lipids (amino acids, peptides, and proteins) ,Arachidonic acid ,Growth inhibition ,Signal transduction ,Signal Transduction - Abstract
Human 15-lipoxygenase (LOX)-2 and mouse 8-LOX represent orthologous members of the LOX family but display different positional specificities and tissue distribution. To study the functional role of 15-LOX-2 and 8-LOX in keratinocytes, an inducible Tet-On gene expression system was established in the premalignant mouse keratinocyte cell line 308. Doxycycline (dox)-induced expression of enzymatically active 15-LOX-2 and 8-LOX led to an inhibition of cell growth that was associated with an inhibition of DNA synthesis, as shown by a 15-46% reduction of 5-bromo-2-deoxy-uridine (BrdU) incorporation. The inhibitory effects were increased in the presence of exogenous arachidonic acid. In contrast, addition of linoleic acid or the LOX inhibitor baicalein reversed the growth-inhibitory effects. Treatment of the cells with 15-hydroxyeicosatetraenoic acid (HETE) or 8-HETE resulted in a similar inhibition of BrdU incorporation, whereas 13-hydroxyoctadecadienoic acid (HODE) and 9-HODE, in contrast, had no effects. Dox-induced keratinocytes showed increased levels of reactive oxygen species (ROS). The antioxidant N-acetyl-L-cysteine and a specific inhibitor of p38 mitogen-activated protein kinase, but not of extracellular signal-regulated kinase 1/2 or c-Jun N-terminal kinase/stress-activated kinases, completely abolished the LOX-induced growth inhibition, indicating a critical role of ROS and p38. Our data suggest that 15-LOX-2 and 8-LOX, although displaying different positional specificity, may use common signaling pathways to induce growth inhibition in premalignant epithelial cells. more...
- Published
- 2007
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30. The Importance of the MM Environment and the Selection of the QM Method in QM/MM Calculations: Applications to Enzymatic Reactions
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Eric André C, Bushnell, Victoria Erica J, Berryman, James W, Gauld, and Russell J, Boyd
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Kinetics ,Catalytic Domain ,Animals ,Anisotropy ,Humans ,Quantum Theory ,Thermodynamics ,Uroporphyrinogen Decarboxylase ,Saccharopine Dehydrogenases ,Molecular Dynamics Simulation ,Anthozoa ,Arachidonate Lipoxygenases ,Substrate Specificity - Abstract
In this chapter, we discuss the influence of an anisotropic protein environment on the reaction mechanisms of saccharopine reductase and uroporphyrinogen decarboxylase, respectively, via the use of a quantum mechanical and molecular mechanical (QM/MM) approach. In addition, we discuss the importance of selecting a suitable DFT functional to be used in a QM/MM study of a key intermediate in the mechanism of 8R-lipoxygenase, a nonheme iron enzyme. In the case of saccharopine reductase, while the enzyme utilizes a substrate-assisted catalytic pathway, it was found that only through treating the polarizing effect of the active site, via the use of an electronic embedding formalism, was agreement with experimental kinetic data obtained. Similarly, in the case of uroporphyrinogen decarboxylase, the effect of the protein environment on the catalytic mechanism was found to be such that the calculated rate-limiting barrier is in good agreement with related experimentally determined values for the first decarboxylation of the substrate. For 8R-lipoxygenase, it was found that the geometries and energies of the multicentered open-shell intermediate complexes formed during the mechanism are quite sensitive to the choice of the density functional theory method. Thus, while density functional theory has become the method of choice in QM/MM studies, care must be taken in the selection of a particular high-level method. more...
- Published
- 2015
31. What are cyclooxygenases and lipoxygenases doing in the driver's seat of carcinogenesis?
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Andreas J. R. Habenicht, K. Müller-Decker, Gerhard Fürstenberger, and P. Krieg
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Lipid Peroxides ,Cancer Research ,Cell Survival ,Linoleic acid ,Lipoxygenase ,Antineoplastic Agents ,Apoptosis ,medicine.disease_cause ,Arachidonate Lipoxygenases ,Linoleic Acid ,chemistry.chemical_compound ,Eicosanoic Acids ,Neoplasms ,medicine ,Animals ,Humans ,Cyclooxygenase Inhibitors ,Neoplasm Invasiveness ,Unsaturated fatty acid ,Cell Proliferation ,chemistry.chemical_classification ,Arachidonic Acid ,Neovascularization, Pathologic ,biology ,Cell growth ,Fatty acid ,Cell Differentiation ,Isoenzymes ,Cell Transformation, Neoplastic ,Oncology ,chemistry ,Eicosanoid ,Biochemistry ,Cyclooxygenase 2 ,Prostaglandin-Endoperoxide Synthases ,Cancer research ,biology.protein ,Arachidonic acid ,Carcinogenesis ,Signal Transduction - Abstract
Substantial evidence supports a functional role for cyclooxygenase- and lipoxygenase-catalyzed arachidonic and linoleic acid metabolism in cancer development. Genetic intervention studies firmly established cause-effect relations for cyclooxygenase-2, but cyclooxygenase-1 may also be involved. In addition, pharmacologic cyclooxygenase inhibition was found to suppress carcinogenesis in both experimental mouse models and several cancers in humans. Arachidonic acid-derived eicosanoid or linoleic acid-derived hydro[peroxy]fatty acid signaling are likely to be involved impacting fundamental biologic phenomena as diverse as cell growth, cell survival, angiogenesis, cell invasion, metastatic potential and immunomodulation. However, long chain unsaturated fatty acid oxidation reactions indicate antipodal functions of distinct lipoxygenase isoforms in carcinogenesis, i.e., the 5- and platelet-type 12-lipoxygenase exhibit procarcinogenic activities, while 15-lipoxygenase-1 and 15-lipoxygenase-2 may suppress carcinogenesis. more...
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- 2006
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32. Regulation of neurotensin receptor function by the arachidonic acid–lipoxygenase pathway in prostate cancer PC3 cells
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Sazzad Hassan, Robert E. Carraway, and David E. Cochrane
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Male ,medicine.medical_specialty ,Inositol Phosphates ,Clinical Biochemistry ,Arachidonate Lipoxygenases ,Binding, Competitive ,chemistry.chemical_compound ,Lipoxygenase ,Cell Line, Tumor ,Internal medicine ,medicine ,Humans ,Receptors, Neurotensin ,Lipoxygenase Inhibitors ,Neurotensin receptor ,Receptor ,Neurotensin ,Unsaturated fatty acid ,Feedback, Physiological ,Leukotriene ,biology ,Prostatic Neoplasms ,Hydrogen Peroxide ,Cell Biology ,Oxidants ,Up-Regulation ,Bombesin receptor ,Endocrinology ,chemistry ,biology.protein ,lipids (amino acids, peptides, and proteins) ,Arachidonic acid ,Reactive Oxygen Species ,Protein Binding ,Signal Transduction - Abstract
Neurotensin (NT) elevates leukotriene levels in animals and stimulates 5-HETE formation in prostate cancer PC3 cells. PC3 cell growth is stimulated by NT and inhibited by lipoxygenase (LOX) blockers. This led us to test LOX blockers (NDGA, MK886, ETYA, Rev5901, AA861 and others) for effects on NT binding and signaling. LOX blockers dramatically enhanced 125 I-neurotensin binding to NT receptor NTR1 in PC3 cells, whereas they inhibited NT-induced inositol phosphate formation. These effects were indirect (binding to isolated membranes was unaffected), receptor-specific (binding to β 2 -adrenergic, V1 a -vasopressin, EGF and bombesin receptor was unaffected) and pathway-specific (cyclooxygenase inhibitors were inactive). NT receptor affinity was increased but receptor number and % internalization were unchanged. Also supporting the involvement of arachidonic acid metabolism in NTR1 regulation was the finding that inhibitors of PLA2 and DAG lipase enhanced NT binding. These findings suggest that NTR1 is regulated by specific feedback mechanism(s) involving lipid peroxidation and/or LOX-dependent processes. more...
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- 2006
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33. Modulation of Cox-1, 5-, 12- and 15-Lox by Popular Herbal Remedies Used in Southern Italy Against Psoriasis and Other Skin Diseases
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Bader, Ammar, Martini, Francesca, Schinella, Guillermo Raúl, Ríos, José Luis, and Prieto, José M.
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Blood Platelets ,Leukotrienes ,Anti-Inflammatory Agents ,Asteraceae ,Arachidonate Lipoxygenases ,Skin Diseases ,Acanthaceae ,Leukocytes ,Animals ,Humans ,Psoriasis ,Research Articles ,Plants, Medicinal ,Plant Extracts ,NF-kappa B ,Rats ,Achillea ,Artemisia ,Italy ,Ciencias Médicas ,Cyclooxygenase 1 ,Prostaglandins ,lipids (amino acids, peptides, and proteins) ,Dermatologic Agents ,Inula ,Herbal medicine ,HeLa Cells ,Phytotherapy - Abstract
Acanthus mollis (Acanthaceae), Achillea ligustica, Artemisia arborescens and Inula viscosa (Asteraceae) are used in Southern Italy against psoriasis and other skin diseases that occur with an imbalanced production of eicosanoids. We here assessed their in vitro effects upon 5-, 12-, 15-LOX and COX-1 enzymes as well as NFκB activation in intact cells as their possible therapeutic targets. All methanol crude extracts inhibited both 5-LOX and COX-1 activities under 200 μg/mL, without significant effects on the 12-LOX pathway or any relevant in vitro free radical scavenging activity. NFκB activation was prevented by all extracts but A. mollis. Interestingly, A. ligustica, A. arborescens and A. mollis increased the biosynthesis of 15(S)-HETE, an anti-inflammatory eicosanoid. A. ligustica (IC50 = 49.5 μg/mL) was superior to Silybum marianum (IC50 = 147.8 μg/mL), which we used as antipsoriatic herbal medicine of reference. Its n-hexane, dichloromethane and ethyl acetate fractions had also inhibitory effects on the LTB4 biosynthesis (IC50s = 9.6, 20.3 and 68 μg/mL, respectively) evidencing that the apolar extracts of A. ligustica are promising active herbal ingredients for future phytotherapeutical products targeting psoriasis., Facultad de Ciencias Médicas more...
- Published
- 2015
34. Arachidonate 8(S)-lipoxygenase
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Gerhard Fürstenberger, Friedrich Marks, and Peter Krieg
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Keratinocytes ,Physiology ,Stratum granulosum ,Biology ,Arachidonate Lipoxygenases ,Biochemistry ,Substrate Specificity ,Exon ,chemistry.chemical_compound ,Lipoxygenase ,Neoplasms ,Hydroxyeicosatetraenoic Acids ,medicine ,Animals ,Humans ,Tissue Distribution ,Gene ,Phylogeny ,Pharmacology ,chemistry.chemical_classification ,ALOX15B ,Cell Differentiation ,Cell Biology ,Hair follicle ,Molecular biology ,Amino acid ,medicine.anatomical_structure ,chemistry ,biology.protein ,Arachidonic acid - Abstract
The recently identified mouse 8( S )-lipoxygenase almost exclusively directs oxygen insertion into the 8( S ) position of arachidonic acid and, with lower efficiency, into the 9( S ) position of linoleic acid. The protein of 677 amino acids displays 78% sequence identity to human 15( S )-lipoxygenase-2 which is considered to be its human orthologue. The 8( S )-lipoxygenase gene, Alox15b, consisting of 14 exons and spanning 14.5 kb is located within a gene cluster of related epidermis-type lipoxygenases at the central region of mouse chromosome 11. 8( S )-Lipoxygenase is predominantly expressed in stratifying epithelia of mice, constitutively in the hair follicle, forestomach, and footsole and inducible in the back skin with strain-dependent variations. The expression is restricted to terminally differentiating keratinocytes, in particular the stratum granulosum and 8( S )-lipoxygenase activity seems to be involved in terminal differentiation of mouse epidermis. Tumor-specific upregulation of 8( S )-lipoxygenase expression and activity indicate a critical role of this enzyme in malignant progression during tumor development in mouse skin. more...
- Published
- 2002
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35. Crystal structure of a lipoxygenase in complex with substrate: the arachidonic acid-binding site of 8R-lipoxygenase
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David B, Neau, Gunes, Bender, William E, Boeglin, Sue G, Bartlett, Alan R, Brash, and Marcia E, Newcomer
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Inflammation ,Models, Molecular ,Arachidonic Acid ,Binding Sites ,integumentary system ,Protein Conformation ,Swine ,Iron ,Crystallography, X-Ray ,Arachidonate Lipoxygenases ,Lipids ,Catalysis ,Oxygen ,Mutagenesis ,Mutation ,Protein Structure and Folding ,Animals ,Humans ,Rabbits ,Protein Binding - Abstract
Lipoxygenases (LOX) play critical roles in mammalian biology in the generation of potent lipid mediators of the inflammatory response; consequently, they are targets for the development of isoform-specific inhibitors. The regio- and stereo-specificity of the oxygenation of polyunsaturated fatty acids by the enzymes is understood in terms of the chemistry, but structural observation of the enzyme-substrate interactions is lacking. Although several LOX crystal structures are available, heretofore the rapid oxygenation of bound substrate has precluded capture of the enzyme-substrate complex, leaving a gap between chemical and structural insights. In this report, we describe the 2.0 Å resolution structure of 8R-LOX in complex with arachidonic acid obtained under anaerobic conditions. Subtle rearrangements, primarily in the side chains of three amino acids, allow binding of arachidonic acid in a catalytically competent conformation. Accompanying experimental work supports a model in which both substrate tethering and cavity depth contribute to positioning the appropriate carbon at the catalytic machinery. more...
- Published
- 2014
36. Discovery of an 11(R)- and 12(S)-lipoxygenase activity in ovaries of the mussel Mytilus edulis
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Elizabeth M. Hill and Gianguido Coffa
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Male ,Ovary ,Biology ,Arachidonate 12-Lipoxygenase ,Arachidonate Lipoxygenases ,Biochemistry ,Gas Chromatography-Mass Spectrometry ,chemistry.chemical_compound ,Testis ,medicine ,Animals ,Chromatography, High Pressure Liquid ,chemistry.chemical_classification ,Organic Chemistry ,Cell Biology ,Eicosapentaenoic acid ,Bivalvia ,Proadifen ,Nordihydroguaiaretic acid ,medicine.anatomical_structure ,Enzyme ,chemistry ,Eicosanoid ,Calcium ,Female ,Arachidonic acid ,Stearidonic acid - Abstract
Eicosanoid biosynthesis was investigated in mussel gonads by incubation of tissue homogenates with radiolabeled arachidonic acid and analysis of the products by radio-high-performance liquid chromatography. No radiolabeled metabolites were formed in homogenates of testes, but two major metabolites were synthesized by ovarian preparations. The radiolabeled metabolites were analyzed by mass spectrometry and chiral chromatography and identified as 11 (R)-hydroxy-5,8,1 2,14-eicosatetraenoic acid and 12(S)-hydroxy-5,8,10,14-eicosatetraenoic acid. In addition, four other nonlabeled metabolites, formed from endogenous substrates, were detected in ovarian extracts. Their structures, determined by mass spectrometric analysis, were the corresponding 11- and 12-hydroxy analogs formed from eicosapentaenoic acid (11-HEPE and 12-HEPE) and 9-hydroxy-6,10,12,15-octadecatetraenoic acid (9-HOTE) and 13-hydroxy-6,9,11,15-octadecatetraenoic acid formecl from stearidonic acid. The biosynthesis of the 11 - and 12-hydroxy products was calcium dependent, localized to the 100,000 x g supernatant cell fraction, and was inhibited by nordihydroguaiaretic acid, but not inhibited by the prostaglandin synthase inhibitors aspirin and indomethacin, or the monoxygenase inhibitor proadifen. Together these data suggested that both the 11 (R)- and 12(S)-hydroxy products were formed from lipoxygenase-type enzymes. Incubation of homogenates of immature ovaries with eicosapentaenoic acid revealed the major product to be I2-HEPE, whereas in mature ovaries mainly 11-HEPE was formed. Extraction of spawned eggs with methanol revealed that predominantly 11-HEPE and 9-HOTE were formed from endogenous substrates. This study shows that female gonads of the mussel express an 11(R)- and 12(S)-lipoxygenase activity whose expression is dependent on differentiation of the ovary. more...
- Published
- 2000
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37. Structural basis for the positional specificity of lipoxygenases
- Author
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Hartmut Kühn
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Hemeproteins ,Models, Molecular ,Physiology ,Lipoxygenase ,Molecular Conformation ,Mutagenesis (molecular biology technique) ,Biology ,Arachidonate Lipoxygenases ,Biochemistry ,Substrate Specificity ,chemistry.chemical_compound ,Animals ,Pharmacology ,chemistry.chemical_classification ,Substrate Interaction ,Binding Sites ,Fatty Acids ,Fatty acid ,Cell Biology ,Enzyme ,Structural biology ,chemistry ,biology.protein ,Arachidonic acid - Abstract
The positional specificity of arachidonic acid oxygenation is currently the decisive parameter for classification of mammalian lipoxygenases but, unfortunately, the structural reasons for lipoxygenase specificity are not well understood. Although there are no direct structural data on lipoxygenase/substrate interaction, experiments with modified fatty acid substrates and mutagenesis studies suggest that for 12- and 15-lipoxygenases, arachidonic acid slides into the substrate-binding pocket with its methyl end ahead. For arachidonate 5- and/or 8-lipoxygenation two alternative models for the enzyme/substrate interaction have been developed: 1) The orientation-determined model and 2) the space-determined model. This review explores the experimental data available on the mechanistic reasons for lipoxygenase specificity and concludes that each of the above-mentioned hypotheses may be valid for arachidonate 5-lipoxygenation under certain circumstances. more...
- Published
- 2000
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38. Identification of Amino Acid Determinants of the Positional Specificity of Mouse 8S-Lipoxygenase and Human 15S-Lipoxygenase-2
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William E. Boeglin, Mitsuo Jisaka, Alan R. Brash, and Richard B. Kim
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Models, Molecular ,endocrine system diseases ,Stereochemistry ,Recombinant Fusion Proteins ,Molecular Sequence Data ,Restriction Mapping ,Crystallography, X-Ray ,Arachidonate Lipoxygenases ,Biochemistry ,Protein Structure, Secondary ,Substrate Specificity ,Mice ,chemistry.chemical_compound ,Residue (chemistry) ,Lipoxygenase ,Escherichia coli ,Aromatic amino acids ,Animals ,Arachidonate 15-Lipoxygenase ,Humans ,Amino Acid Sequence ,Molecular Biology ,Histidine ,DNA Primers ,Mammals ,chemistry.chemical_classification ,Binding Sites ,Base Sequence ,Sequence Homology, Amino Acid ,integumentary system ,biology ,Mutagenesis ,Cell Biology ,Recombinant Proteins ,Amino acid ,Glutamine ,Enzyme ,chemistry ,Mutagenesis, Site-Directed ,biology.protein ,Sequence Alignment - Abstract
Phorbol ester-inducible mouse 8S-lipoxygenase (8-LOX) and its human homologue, 15S-lipoxygenase-2 (15-LOX-2), share 78% identity in amino acid sequences, yet there is no overlap in their positional specificities. In this study, we investigated the determinants of positional specificity using a random chimeragenesis approach in combination with site-directed mutagenesis. Exchange of the C-terminal one-third of the 8-LOX with the corresponding portion of 15-LOX-2 yielded a chimeric enzyme with exclusively 15S-lipoxygenase activity. The critical region was narrowed down to a cluster of five amino acids by expression of multiple cDNAs obtained by in situ chimeragenesis in Escherichia coli. Finally, a pair of amino acids, Tyr603 and His604, was identified as the positional determinant by site-directed mutagenesis. Mutation of both of these amino acids to the corresponding amino acids in 15-LOX-2 (Asp and Val, respectively) converted the positional specificity from 8S to 90% 15S without yielding any other by-products. Mutation of the corresponding residues in 15-LOX-2 to the 8-LOX sequence changed specificity to 50% oxygenation at C-8 for one amino acid substitution and 70% at C-8 for the double mutant. Based on the crystal structure of the reticulocyte 15-LOX, these two amino acids lie opposite the open coordination position of the catalytic iron in a likely site for substrate binding. The change from 8 to 15 specificity entails a switch in the head to tail binding of substrate. Enzymes that react with substrate “head first” (5-LOX and 8-LOX) have a bulky aromatic amino acid and a histidine in these positions, whereas lipoxygenases that accept substrates “tail first” (12-LOX and 15-LOX) have an aliphatic residue with a glutamine or aspartate. Thus, this positional determinant of the 8-LOX and 15-LOX-2 may have significance for other lipoxygenases. more...
- Published
- 2000
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39. Leukotriene A synthase activity of purified mouse skin arachidonate 8-lipoxygenase expressed in Escherichia coli
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Yoshitaka Takahashi, Na Qiao, Hiroyuki Takamatsu, and Tanihiro Yoshimoto
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Leukotrienes ,DNA, Complementary ,Linoleic acid ,Gene Expression ,Biology ,Arachidonate Lipoxygenases ,Substrate Specificity ,Mice ,chemistry.chemical_compound ,Affinity chromatography ,Escherichia coli ,Animals ,Cloning, Molecular ,Molecular Biology ,Skin ,chemistry.chemical_classification ,Arachidonate 5-Lipoxygenase ,Arachidonic Acid ,Leukotriene A4 ,Fatty acid ,Cell Biology ,Molecular biology ,Enzyme ,Biochemistry ,chemistry ,Docosahexaenoic acid ,Arachidonate 5-lipoxygenase ,biology.protein ,Arachidonic acid - Abstract
Mouse skin 8-lipoxygenase was expressed in COS-7 cells by transient transfection of its cDNA in pEF-BOS carrying an elongation factor-1alpha promoter. When crude extract of the transfected COS-7 cells was incubated with arachidonic acid, 8-hydroxy-5,9,11, 14-eicosatetraenoic acid was produced as assessed by reverse- and straight-phase high performance liquid chromatographies. The recombinant enzyme also reacted on alpha-linolenic and docosahexaenoic acids at almost the same rate as that with arachidonic acid. Eicosapentaenoic and gamma-linolenic acids were also oxygenated at 43% and 56% reaction rates of arachidonic acid, respectively. In contrast, linoleic acid was a poor substrate for this enzyme. The 8-lipoxygenase reaction with these fatty acids proceeded almost linearly for 40 min. The 8-lipoxygenase was also expressed in an Escherichia coli system using pQE-32 carrying six histidine residues at N-terminal of the enzyme. The expressed enzyme was purified over 380-fold giving a specific activity of approximately 0.2 micromol/45 min per mg protein by nickel-nitrilotriacetate affinity chromatography. The enzymatic properties of the purified 8-lipoxygenase were essentially the same as those of the enzyme expressed in COS-7 cells. When the purified 8-lipoxygenase was incubated with 5-hydroperoxy-6,8,11, 14-eicosatetraenoic acid, two epimers of 6-trans-leukotriene B4, degradation products of unstable leukotriene A4, were observed upon high performance liquid chromatography. Thus, the 8-lipoxygenase catalyzed synthesis of leukotriene A4 from 5-hydroperoxy fatty acid. Reaction rate of the leukotriene A synthase was approximately 7% of arachidonate 8-lipoxygenation. In contrast to the linear time course of 8-lipoxygenase reaction with arachidonic acid, leukotriene A synthase activity leveled off within 10 min, indicating suicide inactivation. more...
- Published
- 1999
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40. Do lipoxygenases modulate normal or aberrant lympho-hematopoiesis?
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Kenning M. Anderson, Jules E. Harris, and Frank G. Ondrey
- Subjects
Cancer Research ,Programmed cell death ,Arachidonate 5-Lipoxygenase ,Cell growth ,Cellular differentiation ,Hematology ,Biology ,Hematopoietic Stem Cells ,Arachidonate Lipoxygenases ,Hematopoiesis ,Malignant transformation ,Haematopoiesis ,Oncology ,Leukopoiesis ,Gene Targeting ,Immunology ,Cancer research ,Animals ,Lymphopoiesis ,Enzyme Inhibitors ,Stem cell - Abstract
Whether 5- (and the 12- or 15-) lipoxygenases participate in normal or malignantly transformed hematopoietic cell proliferation and differentiation, or contribute to programmed or necrotic cell death has been difficult to decide. Recent evidence concerning these questions is reviewed and some reasons for these difficulties are considered. more...
- Published
- 1999
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41. Affinities of various mammalian arachidonate lipoxygenases and cyclooxygenases for molecular oxygen as substrate
- Author
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Shozo Yamamoto, Ivo Juránek, and Hiroshi Suzuki
- Subjects
Blood Platelets ,Oxygenase ,Swine ,chemistry.chemical_element ,In Vitro Techniques ,Arachidonate 12-Lipoxygenase ,Arachidonate Lipoxygenases ,Oxygen ,Substrate Specificity ,chemistry.chemical_compound ,Lipoxygenase ,Leukocytes ,medicine ,Animals ,Arachidonate 15-Lipoxygenase ,Humans ,Hypoxia ,Molecular Biology ,chemistry.chemical_classification ,Arachidonate 5-Lipoxygenase ,Arachidonic Acid ,biology ,Chemistry ,Membrane Proteins ,Substrate (chemistry) ,Cell Biology ,Hypoxia (medical) ,Affinities ,Recombinant Proteins ,Isoenzymes ,Kinetics ,Enzyme ,Biochemistry ,Cyclooxygenase 2 ,Prostaglandin-Endoperoxide Synthases ,Cyclooxygenase 1 ,biology.protein ,Arachidonic acid ,medicine.symptom - Abstract
In an attempt to study affinities for molecular oxygen of mammalian arachidonate oxygenases, which remain unclarified at present, we determined activities of platelet-type 12-lipoxygenase, leukocyte-type 12-lipoxygenase, 5-lipoxygenase, 15-lipoxygenase, cyclooxygenase-1 and cyclooxygenase-2 at various oxygen concentrations. Activities of all the tested enzymes were assessed by oxygenation of radioactive arachidonic acid under hypoxic conditions, and part of the enzymes were also assayed by monitoring oxygen consumption. Their Km values for oxygen ranged between 10 and 26 μM. These results should be considered in investigations of arachidonic acid metabolism in pathophysiological processes associated with hypoxia. more...
- Published
- 1999
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42. cDNA cloning of a 8-lipoxygenase and a novel epidermis-type lipoxygenase from phorbol ester-treated mouse skin
- Author
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Gerhard Fürstenberger, Friedrich Marks, Andreas Kinzig, Peter Krieg, and Markus Heidt
- Subjects
Lipoxygenase ,Molecular Sequence Data ,Biophysics ,Arachidonate 12-Lipoxygenase ,Arachidonate Lipoxygenases ,Biochemistry ,Mice ,chemistry.chemical_compound ,Endocrinology ,Complementary DNA ,Animals ,Arachidonate 15-Lipoxygenase ,Amino Acid Sequence ,RNA, Messenger ,Cloning, Molecular ,Peptide sequence ,Screening procedures ,Skin ,chemistry.chemical_classification ,Base Sequence ,Sequence Homology, Amino Acid ,Molecular mass ,biology ,Sequence Analysis, DNA ,Molecular biology ,Amino acid ,Molecular Weight ,Open reading frame ,chemistry ,biology.protein ,Phorbol ,Tetradecanoylphorbol Acetate ,Sequence Alignment - Abstract
Using a combination of PCR cloning and conventional screening procedures, we isolated from phorbol ester-treated mouse epidermis two full length cDNA clones encoding novel lipoxygenases. One of the cDNAs turned out to be identical to the recently cloned 8-lipoxygenase [Jisaka et al., J. Biol. Chem. 272 (1997) 24 410–24 416], the open reading frame of the second one corresponded to a protein of 701 amino acids with a calculated molecular mass of 80.6 kDa. The amino acid sequence showed 50.8% identity to human 15-lipoxygenase 2, approximately 40% to 5-lipoxygenase and 35% to 12- and 15-lipoxygenases. A unique structural feature is the insertion of 31 amino acid residues in the amino-terminal part of the molecule. Based on these data, we conclude that this epidermis-derived cDNA encodes a novel lipoxygenase isoform termed provisionally epidermis-type lipoxygenase 2 (e-LOX 2). more...
- Published
- 1998
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43. Molecular Cloning and Functional Expression of a Phorbol Ester-inducible 8S-Lipoxygenase from Mouse Skin
- Author
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Lillian B. Nanney, Richard B. Kim, Alan R. Brash, Mitsuo Jisaka, and William E. Boeglin
- Subjects
DNA, Complementary ,Molecular Sequence Data ,Human skin ,Molecular cloning ,Biology ,Arachidonate Lipoxygenases ,Polymerase Chain Reaction ,Biochemistry ,Gene Expression Regulation, Enzymologic ,Mice ,chemistry.chemical_compound ,Complementary DNA ,Animals ,Humans ,Amino Acid Sequence ,Cloning, Molecular ,Molecular Biology ,Peptide sequence ,Skin ,chemistry.chemical_classification ,Base Sequence ,Sequence Homology, Amino Acid ,integumentary system ,cDNA library ,Cell Biology ,Molecular biology ,Amino acid ,chemistry ,Phorbol ,Tetradecanoylphorbol Acetate ,Arachidonic acid ,HeLa Cells - Abstract
One of the effects of topical application of phorbol ester to mouse skin is the induction of an 8S-lipoxygenase in association with the inflammatory response. Here we report the molecular cloning and characterization of this enzyme. The cDNA was isolated by polymerase chain reaction from mouse epidermis and subsequently from a mouse epidermal cDNA library. The cDNA encodes a protein of 677 amino acids with a calculated molecular mass of 76 kDa. The amino acid sequence has 78% identity to a 15S-lipoxygenase cloned recently from human skin and approximately 40% identity to other mammalian lipoxygenases. When expressed in vaccinia virus-infected Hela cells, the mouse enzyme converts arachidonic acid exclusively to 8S-hydroperoxyeicosatetraenoic acid while linoleic acid is converted to 9S-hydroperoxy-linoleic acid in lower efficiency. Phorbol ester treatment of mouse skin is associated with strong induction of 8S-lipoxygenase mRNA and protein. By Northern analysis, expression of 8S-lipoxygenase mRNA was also detected in brain. Immunohistochemical analysis of phorbol ester-treated mouse skin showed the strongest reaction to 8S-lipoxygenase in the differentiated epidermal layer, the stratum granulosum. The inducibility may be a characteristic feature of the mouse 8S-lipoxygenase and its human 15S-lipoxygenase homologue. more...
- Published
- 1997
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44. Identification of an 8-Lipoxygenase Pathway in Nervous Tissue of Aplysia californica
- Author
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James H. Schwartz, Douglas J. Steel, Tamara L. Tieman, and Steven J. Feinmark
- Subjects
Central Nervous System ,Leukotrienes ,Metabolite ,Arachidonic Acids ,In Vitro Techniques ,Arachidonate 12-Lipoxygenase ,Arachidonate Lipoxygenases ,Biochemistry ,Gas Chromatography-Mass Spectrometry ,chemistry.chemical_compound ,Aplysia ,Hydroxyeicosatetraenoic Acids ,medicine ,Animals ,Molecular Biology ,Chromatography, High Pressure Liquid ,Neurons ,chemistry.chemical_classification ,Arachidonic Acid ,biology ,Nervous tissue ,Cell Biology ,Metabolism ,biology.organism_classification ,Acetylcholine ,Enzyme Activation ,Enzyme ,medicine.anatomical_structure ,chemistry ,Second messenger system ,Arachidonic acid ,Histamine ,Signal Transduction ,medicine.drug - Abstract
Arachidonic acid is converted to (8R)-hydroperoxyeicosa-5,9,11, 14-tetraenoic acid (8-HPETE) during incubations with homogenates of the central nervous system of the marine mollusc, Aplysia californica. 8-HPETE can be reduced to the corresponding hydroxy acid or be enzymatically converted to a newly identified metabolite, 8-ketoeicosa-5,9,11,14-tetraenoic acid (8-KETE). These metabolites were identified by high performance liquid chromatography, UV absorbance, and gas chromatography/mass spectrometry. Stereochemical analysis of the products demonstrate that the neuronal enzyme is an (8R)-lipoxygenase. Previously we have shown that the neurotransmitters, histamine and Phe-Met-Arg-Phe-amide, activate 12-lipoxygenase metabolism in isolated identified Aplysia neurons. We now show that acetylcholine activates the (8R)-lipoxygenase pathway within intact nerve cells. Thus, both (12S)- and (8R)-lipoxygenase co-exist in intact Aplysia nervous tissue but are differentially activated by several neurotransmitters. The precise physiological role of the 8-lipoxygenase products is currently under investigation, but by analogy to the well-described 12-lipoxygenase pathway, we suggest that (8R)-HPETE and 8-KETE may serve as second messengers in Aplysia cholinoceptive neurons. more...
- Published
- 1997
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45. Inhibition of Arachidonate Lipoxygenase Activities by 2-(3,4-Dihydroxyphenyl)ethanol, a Phenolic Compound from Olives
- Author
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Noriko Kohyama, Tadahiro Nagata, Keizo Sekiya, and Shin-ichi Fujimoto
- Subjects
Blood Platelets ,Neutrophils ,Leukotriene B4 ,Arachidonate Lipoxygenases ,Applied Microbiology and Biotechnology ,Biochemistry ,Analytical Chemistry ,chemistry.chemical_compound ,Lipoxygenase ,Oleuropein ,Caffeic acid ,Animals ,Enzyme Inhibitors ,Rats, Wistar ,Molecular Biology ,Cells, Cultured ,Arachidonic Acid ,Molecular Structure ,biology ,Plant Extracts ,Organic Chemistry ,General Medicine ,Phenylethyl Alcohol ,Arachidonate 12-Lipoxygenase ,Rats ,chemistry ,Enzyme inhibitor ,Fruit ,Arachidonate 5-lipoxygenase ,biology.protein ,Arachidonic acid ,Biotechnology - Abstract
The effects of olive fruit extract on arachidonic acid lipoxygenase activities were investigated using rat platelets and rat polymorphonuclear leukocytes (PMNL). Olive extract strongly inhibited both 12-lipoxygenase (12-LO) and 5-lipoxygenase (5-LO) activities. One of the compounds responsible for this inhibition was purified and identified as 2-(3,4-dihydroxyphenyl)ethanol (DPE). DPE inhibited platelet 12-LO activity (IC50, 4.2 microM) and PMNL 5-LO activity (IC50, 13 microM) but not cyclooxygenase activity in cell-free conditions. It also inhibited 12-LO activity in intact platelets (IC50, 50 microM) and reduced leukotriene B4 production in intact PMNL stimulated by A23187 (IC50, 26 microM). The inhibition by DPE of both lipoxygenase activities was stronger than that by oleuropein, caffeic acid, or 7 other related phenolic compounds, especially in intact cells. These results suggest that DPE is a potent specific inhibitor of lipoxygenase activities. more...
- Published
- 1997
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46. Co-regulated expression of glomerular 12/15-lipoxygenase and interleukin-4 mRNAs in rat nephrotoxic nephritis
- Author
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Kamal F. Badr, Fadi G. Lakkis, Naomasa Makita, and Tetsuo Katoh
- Subjects
Kidney Glomerulus ,Molecular Sequence Data ,Biology ,Arachidonate 12-Lipoxygenase ,Arachidonate Lipoxygenases ,Polymerase Chain Reaction ,Proinflammatory cytokine ,Glomerulonephritis ,Gene expression ,medicine ,Animals ,Arachidonate 15-Lipoxygenase ,Humans ,Amino Acid Sequence ,RNA, Messenger ,Cloning, Molecular ,Rats, Wistar ,Interleukin 4 ,DNA Primers ,Regulation of gene expression ,Messenger RNA ,Base Sequence ,Sequence Homology, Amino Acid ,Lymphokine ,medicine.disease ,Molecular biology ,Rats ,Real-time polymerase chain reaction ,Gene Expression Regulation ,Biochemistry ,Nephrology ,Interleukin-4 - Abstract
Co-regulated expression of glomerular 12/15-lipoxygenase and interleukin-4 mRNAs in rat nephrotoxic nephritis. Arachidonate 12- and 15-lipoxygenase (LO) products are generated in experimental glomerulonephritis. 15-S-HETE (a 15-LO product) and lipoxins (interaction products between 5-LO and either 12-LO or 15-LO) counteract the proinflammatory actions of leukotrienes. IL-4 has been shown to up-regulate 15-LO gene expression in human leukocytes. Based on homology with human 15-LO, we cloned a 0.76 kbp fragment of a rat LO cDNA from leukocytes stimulated by interleukin-4 (IL-4). The deduced amino acid sequence shows 71.0% and 60.1% homology to human 15-LO and 12-LO, respectively, and 100% homology to a recently cloned “leukocyte type” rat 12-lipoxygenase enzyme, which possesses significant 15-lipoxygenase activity (heretofore referred to as “12/15-LO”). A deletion mutant was utilized to generate internal standard cRNA in quantitative PCR assays. Glomerular 12/15-LO mRNA increased significantly over controls 24 and 48 hours after NTS injection, then decreased at 72 hours. RNA from NTS glomeruli contained higher levels of 12/15-LO mRNA than that from unstimulated peripheral leukocytes, suggesting that 12/15-LO transcription is up-regulated locally in native and/or infiltrating glomerular cells. Glomerular IL-4 mRNA increased markedly 16 hours post-NTS, and was then reduced, suggesting a potential role for T cell-derived IL-4 in directing the expression of 12/15-LO during glomerulonephritis. This represents the first demonstration of tandem regulated in vivo gene expression for a lymphokine (IL-4) and a lipoxygenase, both of which promote counter-inflammatory influences in immune complex-mediated injury. more...
- Published
- 1994
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47. Regio- and stereochemistry of the dioxygenation reaction catalyzed by (S)-type lipoxygenases or by the cyclooxygenase activity of prostaglandin H synthases
- Author
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Wolf D. Lehmann
- Subjects
Reaction mechanism ,Free Radicals ,Stereochemistry ,Linoleic acid ,Lipoxygenase ,Prostaglandin ,Arachidonate 12-Lipoxygenase ,Arachidonate Lipoxygenases ,Biochemistry ,Substrate Specificity ,Catalysis ,chemistry.chemical_compound ,Physiology (medical) ,Animals ,Arachidonate 15-Lipoxygenase ,Humans ,Mammals ,Arachidonate 5-Lipoxygenase ,Arachidonic Acid ,biology ,Stereoisomerism ,chemistry ,Prostaglandin-Endoperoxide Synthases ,biology.protein ,Substrate specificity ,Arachidonic acid ,Cyclooxygenase - Abstract
Investigations on the regio- and stereochemistry of the reactions of mammalian lipoxygenases and of prostaglandin H synthases are reviewed. The results and concepts are summarized as two reaction box models. The structures of all known (S)-type lipoxygenase products of long-chain fatty acids carrying an all-cis-1,4-diene structural element including mono-, di-, and trihydroxyl products can be accomodated by this model. The model also provides an explanation for leukotriene formation by mammalian lipoxygenases and for the substrate specificity of lipoxygenases towards esterified fatty acids. The reaction box model for the first dioxygenation step of the cyclooxygenase activity of prostaglandin H synthase is stereochemically different from the (S)-type lipoxygenase box model. more...
- Published
- 1994
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48. Inhibition of two-stage skin carcinogenesis as well as complete skin carcinogenesis by oral administration of TMK688, a potent lipoxygenase inhibitor
- Author
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Ryuichi Kato, Hong Jiang, and Satoshi Yamamoto
- Subjects
Cancer Research ,Skin Neoplasms ,Leukotriene B4 ,9,10-Dimethyl-1,2-benzanthracene ,Administration, Topical ,Indomethacin ,Administration, Oral ,DMBA ,Mice, Inbred Strains ,Pharmacology ,Arachidonate Lipoxygenases ,Mice ,chemistry.chemical_compound ,Piperidines ,Benzo(a)pyrene ,Animals ,Anticarcinogenic Agents ,Lipoxygenase Inhibitors ,Anticarcinogen ,Skin ,integumentary system ,biology ,General Medicine ,Biochemistry ,chemistry ,Enzyme inhibitor ,Tetradecanoylphorbol Acetate ,Arachidonate 5-lipoxygenase ,biology.protein ,Female ,Cyclooxygenase - Abstract
1-([5'-(3''-methoxy-4''-ethoxycarbonyloxyphenyl)-2',4'- pentadienoyl]aminoethyl)-4-diphenylmethoxypiperidine (TMK688) is a potent and orally active 5-lipoxygenase inhibitor having anti-histamine activity in its moiety. Recently, we have found that TMK688 also inhibits epidermal cyclooxygenase activity with a potency similar to its inhibiting 5-lipoxygenase. Oral administration of 30 mg/kg TMK688, a dose which markedly inhibits tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA)-stimulated LTB4 formation in mouse skin, markedly inhibited both TPA-promoted and a non-TPA-type tumor promoter anthralin-promoted skin tumor formation in 7,12-dimethylbenz[a]anthracene (DMBA)-initiated CD-1 mice. The inhibitory effect of TMK688 was not due to any damage inflicted on the initiated cells but due to its antitumor-promoting activity. TMK688 not only inhibited two-stage skin carcinogenesis but also inhibited benzo[a]pyrene-caused complete skin carcinogenesis. Throughout the tumorigenesis experiment, the survival rate of animals was 100% and the TMK688-treated mice looked healthy. The body weight gain of TMK688-treated mice was not significantly different from that of non-treated mice. Both TMK688 and 1-([5'-(3''-methoxy-4''-hydroxyphenyl)-2',4'-pentadienoyl]amino eth yl]-4-diphenylmethoxypiperidine (TMK777), an active metabolite of TMK688 having more potent 5-lipoxygenase inhibitory activity and less potent cyclooxygenase inhibitory activity than TMK688, inhibited epidermal 8-lipoxygenase activity induced by a topical application of TPA to mouse skin. The 8-lipoxygenase inhibitory activity of TMK777 was approximately 5 times more potent than that of TMK688. Indomethacin, a typical cylcloxygenase inhibitor, in topical doses which almost completely inhibit epidermal PGE2 formation, failed to inhibit or only slightly inhibited DMBA-initiated and TPA-promoted skin tumor formation. These results suggest that the cyclooxygenase inhibitory effect of TMK688 is not essential for its anti-tumor promoting activity. Although at present a possible contribution of anti-histamine activity cannot be ruled out completely, the anti-tumor promoting action of TMK688 may most probably be related to its anti-lipoxygenase activity. TMK688 seems to be a promising agent for the prevention of skin carcinogenesis. more...
- Published
- 1994
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49. 8R-Lipoxygenase-catalyzed synthesis of a prominent cis-epoxyalcohol from dihomo-γ-linolenic acid: a distinctive transformation compared with S-lipoxygenases
- Author
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Jin K. Cha, Jing Jin, William E. Boeglin, and Alan R. Brash
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Magnetic Resonance Spectroscopy ,Stereochemistry ,Allene ,Lipoxygenase ,Epoxide ,QD415-436 ,Biochemistry ,Arachidonate Lipoxygenases ,Gas Chromatography-Mass Spectrometry ,Substrate Specificity ,chemistry.chemical_compound ,Endocrinology ,8,11,14-Eicosatrienoic Acid ,Sharpless epoxidation ,arachidonic acid ,Animals ,total synthesis ,hepoxilin ,Research Articles ,Dihomo-γ-linolenic acid ,biology ,Plexaura homomalla ,Total synthesis ,Cell Biology ,biology.organism_classification ,Anthozoa ,trans epoxide ,Intramolecular Oxidoreductases ,chemistry ,Hepoxilin ,biology.protein ,Epoxy Compounds ,Arachidonic acid ,18O2 incorporation - Abstract
Conversion of fatty acid hydroperoxides to epoxyalcohols is a well known secondary reaction of lipoxygenases, described for S-specific lipoxygenases forming epoxyalcohols with a trans-epoxide configuration. Here we report on R-specific lipoxygenase synthesis of a cis-epoxyalcohol. Although arachidonic and dihomo-γ-linolenic acids are metabolized by extracts of the Caribbean coral Plexaura homomalla via 8R-lipoxygenase and allene oxide synthase activities, 20:3ω6 forms an additional prominent product, identified using UV, GC-MS, and NMR in comparison to synthetic standards as 8R,9S-cis-epoxy-10S-erythro-hydroxy-eicosa-11Z,14Z-dienoic acid. Both oxygens of (18)O-labeled 8R-hydroperoxide are retained in the product, indicating a hydroperoxide isomerase activity. Recombinant allene oxide synthase formed only allene epoxide from 8R-hydroperoxy-20:3ω6, whereas two different 8R-lipoxygenases selectively produced the epoxyalcohol.A biosynthetic scheme is proposed in which a partial rotation of the reacting intermediate is required to give the observed erythro epoxyalcohol product. This characteristic and the synthesis of cis-epoxy epoxyalcohol may be a feature of R-specific lipoxygenases. more...
- Published
- 2011
50. [Resistance to antiplatelet drugs in patients with cerebrovascular disorders]
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Za, Suslina, Mm, Tanashian, and Ma, Domashenko
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Blood Platelets ,Polymorphism, Genetic ,Ticlopidine ,Aspirin ,Dose-Response Relationship, Drug ,Drug Resistance ,Platelet Activation ,Arachidonate Lipoxygenases ,Clopidogrel ,Secondary Prevention ,Humans ,Patient Compliance ,Intracranial Thrombosis ,Biotransformation ,Platelet Aggregation Inhibitors - Abstract
This review concerns clinical and laboratory resistance to antiplatelet drugs (aspirin and clopidogrel) in patients with cerebrovascular disorders. Results of certain clinical trials showed that laboratory resistance to antiaggregants is associated with recurrent thromboembolic vascular events. The commonest causes of aspirin resistance are production of arachidonic acid metabolites via the lipoxygenase pathway, poor compliance with the treatment, polymorphism of the genes encoding for cyclooxygenase and glycoprotein (GP) IIb/IIIa, endothelial dysfunction. The causes of clopidogrel resistance include inadequate doses of the drug, its low absorption, poor compliance with the treatment, polymorphism of ADP receptors, GP IIb/IIIa and cytochrome P450 genes, acute coronary syndrome and stroke, metabolic syndrome. Therapeutic efficacy of antiaggregants can be improved by increasing their doses, using membranotropic agents, correcting endothelial dysfunction, etc. Because the apparent variability of antiplatelet drug resistance is currently due to the use of different test-systems by different authors, the evaluation of individual sensitivity to a given drug showing laboratory resistance and the choice of alternative therapy are thus far possible only in the framework of clinical studies. Large-scale prospective multicenter trials of antiplatelet drug resistance are needed along with research for better understanding mechanisms of individual platelet sensitivity and resistance to antiaggregants and developing efficacious methods for their correction. more...
- Published
- 2011
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