Evaluation of the risk of foreign compound exposure depends upon a knowledge of xenobiotic metabolism. This proposal targets two enzyme classes, epoxide hydrolases (EHs) and esterases, which are both members of the alpha/beta-hydrolase fold family. Rodent and human EHs and esterases expressed in the baculovirus system are used to develop an understanding of their catalytic mechanism based on a variety of tools including X-ray crystallography, homology modeling, enzyme kinetics, synthesis of enzyme chimera, and site directed mutagenesis. An immediate benefit is the ability to screen human hydrolases for possible inhibitors while a long term goal is development of a mechanistic basis to predict substrate and inhibitor binding. Highly toxic epoxides are hydrolyzed in part by EHs. The structural information gained in objective 1 advances understanding enzyme catalysis and inhibition by several compounds including commercial herbicides and insecticides. The message and gene are being used to determine the basis of the unique dual localization of the sEH in mammalian cells. Objective II addresses the hypothesis that changes in EH activity influence health by changing the metabolic profile of endogenous, biologically active fatty acid epoxides. The hypothesis that biologically activity attributed to epoxides of linoleic acid actually is due to generation of a toxic diol metabolite by the sEH will be tested. Analytic methods for the epoxides, diols and conjugates in serum and urine are being developed based on ion trap GLC-MS and immunoassay. Toxicity and metabolism of these oxidized lipids (oxylipins) are examined in transgenic cells expressing key enzymes, in target cells such as alveolar epithelium and resistant cells such as hepatocytes. These studies will be extended in vivo for evaluating interaction of xenobiotics with oxylipin toxicity. Finally, the hypothesis that oxylipin rations in exposed workers are altered by xenobiotic exposure by monitoring oxylipins and herbicide inhibitors of EH urines will be tested. Inhibition of esterases is a well established cause of toxicity. The esterases primarily responsible for the hydrolysis of the insecticides permethrin and malathion are being purified, cloned and expressed in objective III. The recombinant enzymes will be used for developing a structural basis for predicting inhibitors of these enzymes and thus combinations of insecticides which present toxic risk. For both EHs and esterases the hypothesis of a catalytic tetrad in a two step mechanism will be tested.

Agency
National Institute of Health (NIH)
Institute
National Institute of Environmental Health Sciences (NIEHS)
Type
Method to Extend Research in Time (MERIT) Award (R37)
Project #
5R37ES002710-27
Application #
7256318
Study Section
Special Emphasis Panel (ZRG4-ALTX-1 (01))
Program Officer
Balshaw, David M
Project Start
1980-12-01
Project End
2008-06-30
Budget Start
2007-07-01
Budget End
2008-06-30
Support Year
27
Fiscal Year
2007
Total Cost
$387,806
Indirect Cost
Name
University of California Davis
Department
Zoology
Type
Schools of Earth Sciences/Natur
DUNS #
047120084
City
Davis
State
CA
Country
United States
Zip Code
95618
Harris, Todd R; Kodani, Sean; Rand, Amy A et al. (2018) Celecoxib Does Not Protect against Fibrosis and Inflammation in a Carbon Tetrachloride-Induced Model of Liver Injury. Mol Pharmacol 94:834-841
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Ostermann, Annika I; Herbers, Jan; Willenberg, Ina et al. (2015) Oral treatment of rodents with soluble epoxide hydrolase inhibitor 1-(1-propanoylpiperidin-4-yl)-3-[4-(trifluoromethoxy)phenyl]urea (TPPU): Resulting drug levels and modulation of oxylipin pattern. Prostaglandins Other Lipid Mediat 121:131-7
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