DOCSLIB.ORG

Research Collection

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Research Collection Research Collection Doctoral Thesis Substrate Identification and Functional Analysis of α/β Hydrolase Fold Epoxide Hydrolases Author(s): Hew, Bettina Publication Date: 2019 Permanent Link: https://doi.org/10.3929/ethz-b-000350557 Rights / License: In Copyright - Non-Commercial Use Permitted This page was generated automatically upon download from the ETH Zurich Research Collection. For more information please consult the Terms of use. ETH Library DISS. ETH NO 25662 SUBSTRATE IDENTIFICATION AND FUNCTIONAL ANALYSIS OF α/β HYDROLASE FOLD EPOXIDE HYDROLASES A thesis submitted to attain the degree of DOCTOR OF SCIENCES of ETH ZURICH (Dr. sc. ETH Zurich) presented by BETTINA HEW MSc ETH in Pharmaceutical Sciences, ETH Zurich born on 30.07.1987 citizen of Klosters-Serneus GR accepted on the recommendation of Prof. Dr. Hanns Ulrich Zeilhofer Prof. Dr. Ursula Quitterer und Prof. Dr. Michael Arand 2019 i Preamble PREAMBLE The main project of the present work; “Deciphering the (patho)physiologic role of epoxide hydrolases by ex vivo cartography of their substrate landscape”, was a joined project of me together with Monika Dengler. Originally, it was planned to result in two independent projects after a joined part at the beginning, which did not work out. Thus, the research questions, methods and results were the same for both of us. We worked out the technical details, performed the experiments and analyzed the data with equal contributions. I particularly had the lead in AAV production and cell culture work, while M. Dengler had the lead in LC-MS/MS method development and analysis. In the two corresponding, individually written PhD dissertations the experimental parts were divided accordingly. For the sake of completeness missing sections were summarized and referred to the respective other work. In contrast, the side project dealt with in this work; “The CYP blocker MS-PPOH is an inhibitor of sEH”, was my individual project. i Summary SUMMARY Epoxide hydrolases (EHs) are enzymes that convert epoxides to their corresponding vicinal diols. In mammals, they play a central role in the detoxification of genotoxic epoxides and have an important function in the regulation of physiological processes by hydrolyzing signaling epoxides. The best characterized EHs are the microsomal EH (mEH) and the soluble EH (sEH). mEH is generally considered to be primarily important in detoxification and only recently gained attention in signaling molecule metabolism. sEH, on the other hand, has long been recognized as the key enzyme in the hydrolysis of signaling fatty acid epoxides (FAEs), such as epoxyeicosatrienoic acids (EETs). The most recently discovered family member, EH3, was also shown to hydrolyze FAEs, thus a role in mammalian physiology seems likely. EH4 and mesoderm specific transcript (MEST) are also predicted to be mammalian EHs based on their primary structure, but their EH activity is not yet proven. Bacteria also possess EHs, sometimes with pathophysiologic consequences for host organisms as reported for the CFTR inhibitory factor (Cif), secreted by P. aeruginosa. However, the exact mechanism by which Cif fulfills its pathologic function is still unknown. The knowledge about the aforementioned EHs is typically based on in vitro analysis of their substrate preference and does not allow direct conclusions on their real role in mammalian physiology. Thus, the general aim of the main project of this work was to identify the major (patho)physiologic roles of the six EHs via the identification of their preferred in vivo substrates. For this purpose, a novel unbiased in vivo substrate trapping approach, taking advantage of the characteristic molecular mechanism of EHs, was developed and applied. EH variants able to trap their substrate in a covalent ester-bond due to introduction of a point mutation, were expressed in target tissues of mice using adeno-associated virus (AAV) infection. The resulting enzyme-substrate complex was subsequently retrieved from tissue lysates by His-tag based affinity enrichment and analyzed using liquid chromatography-tandem mass spectrometry (LC-MS/MS) to identify the bound substrates. A LC-MS/MS method to analyze bound substrates was successfully developed. The method could be confirmed by in vitro trapping experiments with sEH trapping mutant and its known substrate 14,15- EET. Moreover, successful AAV-mediated trapping EH expression was detected in different target organs of mice by Western blot and immunohistochemistry (IHC) stainings. However, in a comprehensive series of experiments, it was not possible to identify or verify any substrates with this novel trapping approach. The primary issues were the expression of non-functional trapping mutants unable to bind their substrate (i.e. mEH, MEST, Cif) and the selective loss of the peptides during sample ii Summary preparation for the LC-MS/MS analysis (i.e. sEH, EH3, EH4). Another shortcoming was that the selected AAV serotypes did not lead to transgene expression in the relevant organs for certain EHs (i.e. Cif, EH3). In addition, transgene expression for some EH trapping mutants was low and thus hindered their analysis (i.e. EH3, EH4). Finally, further optimization is required for the success of our developed trapping method. Moreover, trapping attempts in vitro could be a valuable alternative for Cif and EH3 substrate identification. Unexpectedly, mice expressing the mEH trapping mutant in the brain developed a striking progressive trembling phenotype. This phenotype was not observed in any other group of AAV-infected mice. A potential underlying cause is a loss of dopaminergic neurons in the brain motor system of these mice, producing a pathology similar to Parkinson’s disease. However, the exact mechanism remains unclear and needs further examination. In a smaller side project pursued in parallel to the main project, a classic in vitro inhibitor screening was performed. The cytochrome P450 (CYP) inhibitor N-(methylsulfonyl)-2-(2-propynyloxy)- benzenehexanamide (MS-PPOH), which is generally used in vitro and in vivo to prevent EET formation, was examined for its potential to inhibit mEH and sEH activity. While it did not inhibit mEH, MS-PPOH inhibition of sEH was almost two orders of magnitude stronger compared to CYPs. This would lead to erroneous conclusions concerning the physiological EET functions when sEH inhibition is not considered in experiments using MS-PPOH. Nonetheless, additional studies to examine the consequences of these findings in vivo would be of major importance. In the present work, novel insight into EH physiology has been obtained and a method has been established which has the potential to enable unbiased ex vivo substrate identification for EHs in the near future. iii Zusammenfassung ZUSAMMENFASSUNG Epoxidhydrolasen (EHs) sind Enzyme, die Epoxide zu ihren entsprechenden Diolen hydrolysieren. Sie spielen eine zentrale Rolle in der Entgiftung genotoxischer Epoxide und haben eine wichtige Funktion in der Regulation physiologischer Prozesse, indem sie Signalmoleküle hydrolysieren. Die bestcharakterisierten Säugetier-EHs sind die mikrosomale EH (mEH) und die lösliche EH (sEH). Die mEH wird hauptsächlich im Fremdstoffmetabolismus angesiedelt und erlangte erst vor Kurzem Aufmerksamkeit für ihre Funktion im Signalmolekülstoffwechsel. Die sEH hingegen ist das Schlüsselenzym im Umsatz von Fettsäureepoxiden (FAEs), wie Epoxyeicosatriensäuren (EETs). Auch für die erst kürzlich entdeckte EH3 wurde gezeigt, dass sie FAEs hydrolysiert, was eine physiologische Aufgabe nahelegt. Basierend auf ihren Primärstrukturen, sind auch EH4 und mesoderm specific transcript (MEST) potentielle Säugetier-EHs. Ihre EH Aktivität ist jedoch noch nicht erwiesen. Bakterien besitzen auch EHs, die pathophysiologische Konsequenzen für den Wirt mit sich ziehen können. Ein Beispiel dafür ist CFTR inhibitory factor (Cif), ein Virulenzfaktor sekretiert von P. aeruginosa. Der genaue Mechanismus, mit welchem Cif seine pathologische Funktion erfüllt, ist aber noch ungeklärt. Das Wissen über die genannten EHs basiert typischerweise auf in vitro Untersuchen ihrer Substratpräferenz und erlaubt darum keine direkten Rückschlüsse auf ihre eigentliche physiologische Aufgabe. Das Ziel des Hauptprojekts dieser Arbeit war deshalb die Aufklärung der jeweiligen (patho)physiologischen Hauptaufgaben der sechs EHs, mittels Identifizierung ihrer bevorzugten in vivo Substrate. Dazu wurde eine neuartige, unvoreingenommene in vivo Substrat-Fangmethode entwickelt und angewendet. EH Varianten, die aufgrund einer Punktmutation ihr Substrat in einer kovalenten Esterbindung irreversibel festhalten (Trapping-Mutanten), wurden über Adeno-assoziierte Viren (AAV) in Zielorganen von Mäusen exprimiert. Die Enzym-Substrat Komplexe wurden anschliessend via His- Tag Affinitätschromatographie aus dem Gewebe angereichert und mit Flüssigchromatographie- Tandem Massenspektrometrie (LC-MS/MS) analysiert, um gebundene Substrate zu identifizieren. Eine LC-MS/MS Methode für den Nachweis gebundener Substrate wurde erfolgreich entwickelt. Die Methode konnte durch in vitro Einfangexperimente mit Trapping-sEH und ihrem bekannten Substrat 14,15-EET bestätigt werden. Zudem konnte die AAV vermittelte Expression von Trapping-EHs in verschiedenen Zielorganen von Mäusen mittels Westernblot und immunohistochemischen (IHC) Färbungen nachgewiesen werden. Nichtsdestotrotz war es in einer umfangreichen Serie von Experimenten nicht möglich mit der neuartigen Methode neue Substrate zu identifizieren oder bekannte Substrate zu verifizieren.
Load more

Recommended publications
  • Role of Epoxide Hydrolases in Lipid Metabolism
    Biochimie 95 (2013) 91e95 Contents lists available at SciVerse ScienceDirect Biochimie journal homepage: www.elsevier.com/locate/biochi Mini-review Role of epoxide hydrolases in lipid metabolism Christophe Morisseau* Department of Entomology and U.C.D. Comprehensive Cancer Center, One Shields Avenue, University of California, Davis, CA 95616, USA article info abstract Article history: Epoxide hydrolases (EH), enzymes present in all living organisms, transform epoxide-containing lipids to Received 29 March 2012 1,2-diols by the addition of a molecule of water. Many of these oxygenated lipid substrates have potent Accepted 8 June 2012 biological activities: host defense, control of development, regulation of blood pressure, inflammation, Available online 18 June 2012 and pain. In general, the bioactivity of these natural epoxides is significantly reduced upon metabolism to diols. Thus, through the regulation of the titer of lipid epoxides, EHs have important and diverse bio- Keywords: logical roles with profound effects on the physiological state of the host organism. This review will Epoxide hydrolase discuss the biological activity of key lipid epoxides in mammals. In addition, the use of EH specific Epoxy-fatty acids Cholesterol epoxide inhibitors will be highlighted as possible therapeutic disease interventions. Ó Juvenile hormone 2012 Elsevier Masson SAS. All rights reserved. 1. Introduction hydrolyzed by a water molecule [8]. Based on this mechanism, transition-state inhibitors of EHs have been designed (Fig. 1B). Epoxides are three atom cyclic ethers formed by the oxidation of These ureas and amides are tight-binding competitive inhibitors olefins. Because of their highly polarized oxygen-carbon bonds and with low nanomolar dissociation constants (KI) [9] [10].
    [Show full text]
  • Disruption of Soluble Epoxide Hydrolase Dimerization As a Novel Therapeutic Target for Stroke
    DISRUPTION OF SOLUBLE EPOXIDE HYDROLASE DIMERIZATION AS A NOVEL THERAPEUTIC TARGET FOR STROKE By Jonathan W. Nelson A THESIS/DISSERTATION Presented to the Department of Molecular and Medical Genetics and the Oregon Health & Science University School of Medicine in partial fulfillment of the requirements for the degree of Doctor of Philosophy April 2013 TABLE OF CONTENTS List of Figures ...................................................................................................................... vi List of Tables ...................................................................................................................... vii List of Abbreviations ......................................................................................................... viii Acknowledgements ........................................................................................................... xiii Dissertation Abstract ........................................................................................................ xvi Chapter 1 Targeting Soluble Epoxide Hydrolase Dimerization As a Stroke Therapy ......... 1 Abstract ........................................................................................................................... 1 Stroke and Society ........................................................................................................... 2 Prevention and Treatment of Stroke .............................................................................. 3 Soluble Epoxide Hydrolase and Stroke ..........................................................................
    [Show full text]
  • Analysis of Soybean Embryonic Axis Proteins by Two-Dimensional Gel Electrophoresis and Mass Spectrometry
    Journal of Basic & Applied Sciences, 2013, 9, 309-332 309 Analysis of Soybean Embryonic Axis Proteins by Two-Dimensional Gel Electrophoresis and Mass Spectrometry Savithiry S. Natarajana,*, Hari B. Krishnanb, Farooq Khana, Xi Chena, Wesley M. Garrettc and Dilip Lakshmand aUSDA-ARS, Soybean Genomics and Improvement Laboratory, Beltsville, MD 20705, USA bUSDA-ARS, Plant Genetics Research Unit, University of Missouri, Columbia, MO 65211, USA cUSDA-ARS, Animal Science and Biotechnology Laboratory, Beltsville, MD 20705, USA dUSDA-ARS, Sustainable Agricultural Systems Laboratory, Beltsville, MD 20705, USA Abstract: A proteomic approach based on two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) for protein separation and subsequent mass spectrometry (MS) for protein identification was applied to establish a proteomic reference map for the soybean embryonic axis. Proteins were extracted from dissected embryonic axes and separated in the first dimension using a pH range from 4-7. A total of 401 protein spots were isolated, digested with trypsin, and analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS). We identified 335 protein spots by searching National Center for Biotechnology Information (NCBI) non redundant databases using the Mascot search engine and found a total of 200 unique proteins. Gene Ontology (GO) analysis was employed to understand the molecular processes in which the identified embryonic axes proteins are involved. The majority of proteins play a functional role in catalytic activity (42.9%) and binding (39.3%), followed by nutrient reservoir activity (5.3%), structural molecular activity (4.0%), antioxidant activity (3.2%), transporter activity (2.4%), enzyme regulator activity (1.2%), molecular transducer activity (0.8%), and transcription regulator activity (0.8%).
    [Show full text]
  • Genomic Platforms and Molecular Physiology of Insect Stress Tolerance
    Genomic Platforms and Molecular Physiology of Insect Stress Tolerance DISSERTATION Presented in Partial Fulfillment of the Requirements for the Degree Doctor of Philosophy in the Graduate School of The Ohio State University By Justin Peyton MS Graduate Program in Evolution, Ecology and Organismal Biology The Ohio State University 2015 Dissertation Committee: Professor David L. Denlinger Advisor Professor Zakee L. Sabree Professor Amanda A. Simcox Professor Joseph B. Williams Copyright by Justin Tyler Peyton 2015 Abstract As ectotherms with high surface area to volume ratio, insects are particularly susceptible to desiccation and low temperature stress. In this dissertation, I examine the molecular underpinnings of two facets of these stresses: rapid cold hardening and cryoprotective dehydration. Rapid cold hardening (RCH) is an insect’s ability to prepare for cold stress when that stress is preceded by an intermediate temperature for minutes to hours. In order to gain a better understanding of cold shock, recovery from cold shock, and RCH in Sarcophaga bullata I examine the transcriptome with microarray and the metabolome with gas chromatography coupled with mass spectrometry (GCMS) in response to these treatments. I found that RCH has very little effect on the transcriptome, but results in a shift from aerobic metabolism to glycolysis/gluconeogenesis during RCH and preserved metabolic homeostasis during recovery. In cryoprotective dehydration (CD), a moisture gradient is established between external ice and the moisture in the body of an insect. As temperatures decline, the external ice crystals grow, drawing in more moisture which dehydrates the insect causing its melting point to track the ambient temperature. To gain a better understanding of CD and dehydration in Belgica antarctica I explore the transcriptome with RNA sequencing ii and the metabolome with GCMS.
    [Show full text]
  • Division of Intramural Research NAEHS Council Update September 2021
    Division of Intramural Research NAEHS Council Update September 2021 DIR RECRUITMENTS Tenure-Track Investigator in the Clinical Research Branch The National Institute of Environmental Health Sciences (NIEHS) is recruiting outstanding candidates for a Tenure-Track Investigator position in the Clinical Research Branch within the Division of Intramural Research at the NIEHS in Research Triangle Park, NC. The individual selected for this position will have a strong record of participation and publications in patient- oriented research, defined as research that requires direct interactions with human subjects and may include the development of new technologies, understanding mechanisms of human disease, therapeutic interventions and/or clinical trials. The Clinical Research Branch is interested in candidates with expertise in areas such as endocrinology, neuroendocrinology, metabolism, exercise, sleep, immune-mediated diseases, pulmonology, and human genetics, among others. Applicants should have an M.D. or equivalent doctoral degree with direct clinical research experience, with three or more years of research training in clinical research and publications and other evidence of the ability to design and carry out original, innovative patient-oriented research. Dr. Alison Motsinger-Reif, Chief of the Biostatistics and Computational Biology Branch serves as chair of the search committee which was launched on July 8, 2019. This recruitment has been temporarily placed on hold. Medical Director of the Clinical Research Unit The Division of Intramural Research is seeking an accomplished physician scientist to serve as Senior Clinician in the Clinical Research Branch and Medical Director of the Clinical Research Unit (CRU), a stand-alone facility that sees over 1,000 patients and research participants annually with a budget of over $3M.
    [Show full text]
  • EST and Microarray Analysis of Horn Development in Onthophagus Beetles Teiya Kijimoto1, James Costello1,2, Zuojian Tang3, Armin P Moczek1 and Justen Andrews*1
    BMC Genomics BioMed Central Research article Open Access EST and microarray analysis of horn development in Onthophagus beetles Teiya Kijimoto1, James Costello1,2, Zuojian Tang3, Armin P Moczek1 and Justen Andrews*1 Address: 1Department of Biology, Indiana University, Bloomington, Indiana, 47405, USA, 2School of Informatics, Indiana University, Bloomington, Indiana, 47405, USA and 3Center for Genomics and Bioinformatics, Indiana University, Bloomington, Indiana, 47405, USA Email: Teiya Kijimoto - [email protected]; James Costello - [email protected]; Zuojian Tang - [email protected]; Armin P Moczek - [email protected]; Justen Andrews* - [email protected] * Corresponding author Published: 30 October 2009 Received: 23 March 2009 Accepted: 30 October 2009 BMC Genomics 2009, 10:504 doi:10.1186/1471-2164-10-504 This article is available from: http://www.biomedcentral.com/1471-2164/10/504 © 2009 Kijimoto et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Abstract Background: The origin of novel traits and their subsequent diversification represent central themes in evo-devo and evolutionary ecology. Here we explore the genetic and genomic basis of a class of traits that is both novel and highly diverse, in a group of organisms that is ecologically complex and experimentally tractable: horned beetles. Results: We developed two high quality, normalized cDNA libraries for larval and pupal Onthophagus taurus and sequenced 3,488 ESTs that assembled into 451 contigs and 2,330 singletons.
    [Show full text]
  • Identify Epoxide Hydrolase Substrates in Vivo
    Zurich Open Repository and Archive University of Zurich Main Library Strickhofstrasse 39 CH-8057 Zurich www.zora.uzh.ch Year: 2019 Trapped: Developing an Unbiased Approach to Identify Epoxide Hydrolase Substrates in Vivo Dengler, Monika Abstract: Epoxide hydrolases (EHs) of the ฀/฀ hydrolase fold enzyme family hydrolyze epoxides to the corresponding vicinal diols. In mammals, epoxides are mainly formed within the body through epoxi- dation of xenobiotic or endogenous substrates by cytochrome P450-dependent monooxygenases (CYPs). Two of the five known mammalian EHs are well characterized. The microsomal epoxide hydrolase (mEH) is primarily involved in detoxification of carcinogenic epoxides derived from xenobiotic compounds and the soluble epoxide hydrolase (sEH) primarily regulates endogenous signaling epoxides derived from fatty acids. Although some substrates are known for the epoxide hydrolase 3 (EH3), its function remains unclear. For the epoxide hydrolase 4 (EH4) and mesoderm specific transcript (MEST) no substrates are known, but knockout of MEST in mice leads to growth retardation and a behavioral phenotype. Another EH with activity in the human body is the CFTR inhibitory factor (Cif), a virulence factor secreted by the opportunistic pathogen Pseudomonas aeruginosa. Cif reduces the surface expression of the chloride chan- nel CFTR in human airway endothelial cells, resulting in increased mucus viscosity facilitating bacterial colonization. The enzyme activity of Cif is crucial for the effect on CFTR but the molecular target re- mains to be identified. In this project, we aimed to identify the physiologically relevant substrate(s) ofthe aforementioned EHs. For the bacterial EH Cif, a substrate screening was performed using recombinantly expressed enzyme, but kinetic analysis suggested that the found substrates 14,15-epoxyeicosatrienoic acid (14,15-EET), 17,18-epoxyeicosatetraenoic acid, and 19,20-epoxydocosapentaenoic acid are not relevant in vivo.
    [Show full text]
  • Thesis Martina Decker, Characterization EH3 & EH4
    Zurich Open Repository and Archive University of Zurich Main Library Strickhofstrasse 39 CH-8057 Zurich www.zora.uzh.ch Year: 2010 Expression and characterization of the novel human epoxide hydrolases EH3 and EH4 Decker, Martina Marion Elisabeth Posted at the Zurich Open Repository and Archive, University of Zurich ZORA URL: https://doi.org/10.5167/uzh-46370 Dissertation Published Version Originally published at: Decker, Martina Marion Elisabeth. Expression and characterization of the novel human epoxide hydro- lases EH3 and EH4. 2010, University of Zurich, Faculty of Science. EXPRESSION AND CHARACTERIZATION OF THE NOVEL HUMAN EPOXIDE HYDROLASES EH3 AND EH4 DISSERTATION zur Erlangung der naturwissenschaftlichen Doktorwürde (Dr. sc. nat.) vorgelegt der Mathematisch-naturwissenschaftlichen Fakultät der Universität Zürich von Martina Marion Elisabeth Decker aus Deutschland Promotionskomitee Prof. Dr. Michael Arand Prof. Dr. Jean-Marc Fritschy Prof. Dr. Thierry Hennet Zürich, 2010 2 3 4 ACKNOWLEDGEMENTS First, I would like to thank Michael Arand for supervising me and giving me the opportunity to do my PhD in his group. Many thanks for sending me to San Diego and Baltimore (the best steak ever) and of course for the intensive proofreading and his help during writing this thesis. I would also like to thank “Miss Annette” Cronin for simply everything. For helping me with any problems in the lab and outside the lab, for answering all my questions on LC-MS/MS and protein purification and for wonderful barbecues with wonderful potato salad. It was great going to Las Vegas and Baltimore with you! I am really going to miss you. Also a big thank you to Magdalena Adamska, for introducing me into the secrets of molecular biology and for granting me asylum in the very beginning of my PhD.
    [Show full text]
  • Download from the ETH Zurich Research Collection
    Zurich Open Repository and Archive University of Zurich Main Library Strickhofstrasse 39 CH-8057 Zurich www.zora.uzh.ch Year: 2019 Substrate Identification and Functional Analysis of ฀/฀ Hydrolase Fold Epoxide Hydrolases Hew, Bettina DOI: https://doi.org/10.3929/ethz-b-000350557 Posted at the Zurich Open Repository and Archive, University of Zurich ZORA URL: https://doi.org/10.5167/uzh-186648 Dissertation Published Version Originally published at: Hew, Bettina. Substrate Identification and Functional Analysis of ฀/฀ Hydrolase Fold Epoxide Hydrolases. 2019, ETH Zurich. DOI: https://doi.org/10.3929/ethz-b-000350557 Research Collection Doctoral Thesis Substrate Identification and Functional Analysis of α/β Hydrolase Fold Epoxide Hydrolases Author(s): Hew, Bettina Publication Date: 2019 Permanent Link: https://doi.org/10.3929/ethz-b-000350557 Rights / License: In Copyright - Non-Commercial Use Permitted This page was generated automatically upon download from the ETH Zurich Research Collection. For more information please consult the Terms of use. ETH Library DISS. ETH NO 25662 SUBSTRATE IDENTIFICATION AND FUNCTIONAL ANALYSIS OF α/β HYDROLASE FOLD EPOXIDE HYDROLASES A thesis submitted to attain the degree of DOCTOR OF SCIENCES of ETH ZURICH (Dr. sc. ETH Zurich) presented by BETTINA HEW MSc ETH in Pharmaceutical Sciences, ETH Zurich born on 30.07.1987 citizen of Klosters-Serneus GR accepted on the recommendation of Prof. Dr. Hanns Ulrich Zeilhofer Prof. Dr. Ursula Quitterer und Prof. Dr. Michael Arand 2019 i Preamble PREAMBLE The main project of the present work; “Deciphering the (patho)physiologic role of epoxide hydrolases by ex vivo cartography of their substrate landscape”, was a joined project of me together with Monika Dengler.
    [Show full text]
  • The Multifaceted Role of Epoxide Hydrolases in Human Health and Disease
    International Journal of Molecular Sciences Review The Multifaceted Role of Epoxide Hydrolases in Human Health and Disease Jérémie Gautheron 1,2 and Isabelle Jéru 1,2,3,* 1 Centre de Recherche Saint-Antoine (CRSA), Inserm UMRS_938, Sorbonne Université, 75012 Paris, France; [email protected] 2 Institute of Cardiometabolism and Nutrition (ICAN), Hôpital Pitié-Salpêtrière, Assistance Publique-Hôpitaux de Paris, 75013 Paris, France 3 Laboratoire Commun de Biologie et Génétique Moléculaires, Hôpital Saint-Antoine, Assistance Publique-Hôpitaux de Paris, 75012 Paris, France * Correspondence: [email protected] Abstract: Epoxide hydrolases (EHs) are key enzymes involved in the detoxification of xenobiotics and biotransformation of endogenous epoxides. They catalyze the hydrolysis of highly reactive epoxides to less reactive diols. EHs thereby orchestrate crucial signaling pathways for cell homeostasis. The EH family comprises 5 proteins and 2 candidate members, for which the corresponding genes are not yet identified. Although the first EHs were identified more than 30 years ago, the full spectrum of their substrates and associated biological functions remain partly unknown. The two best-known EHs are EPHX1 and EPHX2. Their wide expression pattern and multiple functions led to the development of specific inhibitors. This review summarizes the most important points regarding the current knowledge on this protein family and highlights the particularities of each EH. These different enzymes can be distinguished by their expression pattern, spectrum of associated substrates, sub-cellular localization, and enzymatic characteristics. We also reevaluated the pathogenicity of previously reported variants in genes that encode EHs and are involved in multiple disorders, in light of large datasets that were made available due to the broad development of next generation sequencing.
    [Show full text]
  • Mammalian Epoxide Hydrolases in Xenobiotic Metabolism and Signalling
    Arch Toxicol (2009) 83:297–318 DOI 10.1007/s00204-009-0416-0 REVIEW ARTICLE Mammalian epoxide hydrolases in xenobiotic metabolism and signalling Martina Decker · Michael Arand · Annette Cronin Received: 11 February 2009 / Accepted: 16 February 2009 / Published online: 2 April 2009 © Springer-Verlag 2009 Abstract Epoxide hydrolases catalyse the hydrolysis of in the detoxiWcation of genotoxic epoxides and (2) have an electrophilic—and therefore potentially genotoxic—epox- important function in the regulation of physiological pro- ides to the corresponding less reactive vicinal diols, which cesses by the control of signalling molecules with an epox- explains the classiWcation of epoxide hydrolases as typical ide structure. detoxifying enzymes. The best example is mammalian microsomal epoxide hydrolase (mEH)—an enzyme prone Keywords Epoxide hydrolase · Xenobiotic metabolism · to detoxiWcation—due to a high expression level in the EPHX · ABHD · Lipid signalling · peg1/MEST · EET · liver, a broad substrate selectivity, as well as inducibility by Cholesterol · Lipid phosphatase foreign compounds. The mEH is capable of inactivating a large number of structurally diVerent, highly reactive epox- ides and hence is an important part of the enzymatic Introduction defence of our organism against adverse eVects of foreign compounds. Furthermore, evidence is accumulating that Epoxide hydrolases (EC 3.3.2.7-11) catalyse the hydrolysis mammalian epoxide hydrolases play physiological roles of epoxides to the corresponding vicinal diols by the addi- other than detoxiWcation, particularly through involvement tion of water. Xenobiotic derived epoxides can be formed in signalling processes. This certainly holds true for soluble within the body as reactive intermediates during metabolic epoxide hydrolase (sEH) whose main function seems to be processes by monooxygenation of carbon double bonds in the turnover of lipid derived epoxides, which are signalling oleWnes or aromatic ring systems.
    [Show full text]
  • "Mineração De Genes Em Regiões Genômicas Bovinas Associadas À Resistência Ao Carrapato Rhipicephalus (Boophilus) Microplus"
    Programa de Pós Graduação em Genética Evolutiva e Biologia Molecular Universidade Federal de São Carlos Vitor Catoia "Mineração de genes em regiões genômicas bovinas associadas à resistência ao carrapato Rhipicephalus (Boophilus) microplus" São Carlos 2014 VITOR CATOIA (Biólogo) "Mineração de genes em regiões genômicas bovinas associadas à resistência ao carrapato Rhipicephalus (Boophilus) microplus" Dissertação de mestrado apresentada ao Programa de Pós-Graduação em Genética Evolutiva e Biologia Molecular do Centro de Ciências Biológicas e da Saúde da Universidade Federal de São Carlos, como parte dos requisitos para obtenção do título de Mestre em Genética Evolutiva e Biologia Molecular. Orientadora: Dra. Luciana Correia de Almeida Regitano Coorientadora: Dra. Daniela Dantas Moré São Carlos 2014 Ficha catalográfica elaborada pelo DePT da Biblioteca Comunitária da UFSCar Catoia, Vitor. C366mg Mineração de genes em regiões genômicas bovinas associadas à resistência ao carrapato Rhipicephalus (Boophilus) microplus / Vitor Catoia. -- São Carlos : UFSCar, 2015. 91 f. Dissertação (Mestrado) -- Universidade Federal de São Carlos, 2014. 1. Genética. 2. Animais - melhoramento genético. 3. Infestação. I. Título. CDD: 575 (20a) A Deus, Ao meu querido tio Adilson Pulgrossi (in memorian) Aos meus pais Idelma e Renato, Dedico AGRADECIMENTOS A Deus, que em todo momento me iluminou, me guiou deu forças para enfrentar todas as dificuldades durante minha trajetória. À minha orientadora Dra. Luciana Correia de Almeida Regitano, pelos ensinamentos, orientação, paciência e por me conceder a oportunidade de realizar meu projeto na Embrapa Pecuária Sudeste. À minha coorientadora Dra. Daniela Dantas Moré, pelo apoio, paciência e transmissão dos conhecimentos. Ao Dr. Maurício de Alvarenga Mudadu, pela disponibilidade e ajuda nas análises computacionais.
    [Show full text]