Soluble epoxide hydrolase in rat inflammatory cells is indistinguishable from soluble epoxide hydrolase in rat liver.
Soluble epoxide hydrolase (sEH) is a ubiquitous mammalian enzyme for which liver and kidney are reported to have the highest activity. We have shown that the soluble epoxide hydrolase (sEH) activity present in rat neutrophils and macrophages is kinetically, immunologically, and physically indistinguishable from rat liver cytosolic sEH. Cytosol from rat liver or inflammatory cells and recombinant rat sEH were incubated with trans-diphenylpropene oxide (tDPPO), a selective substrate for sEH. The tDPPO hydration activity we observed in inflammatory cell cytosol was lower than that from liver. The Km for tDPPO hydration observed in rat inflammatory cell cytosol was the same as the Km for rat liver cytosol (10 microM). Recombinant rat sEH and cytosol from rat liver or inflammatory cells were incubated with the sEH inhibitors, chalcone oxide, 4-fluorochalcone oxide, and 4-phenylchalcone oxide. The IC50 values were 40, 8, and 0.4 microM, respectively, in all samples. Furthermore, sEH activity could be completely immunoprecipitated out of the samples, and the amount of antibody required to do so was apparently identical, regardless of the source of enzyme. SDS-polyacrylamide gel electrophoresis followed by Western blot analysis revealed a single sEH band with a molecular weight of 62 kDa. Isoelectric focusing followed by Western blot analysis revealed multiple bands containing tDPPO-hydrating activity. Although the inflammatory cell bands had the same pattern as those from liver cytosol, the recombinant sEH showed a different banding pattern. These multiple bands were not an artifact of the IEF gel selected. Furthermore, in a 2-dimensional IEF gel, the bands re-migrated to the same position. The presence of sEH in inflammatory cells suggests that this enzyme may have an important endogenous function.
Draper
A J
AJ
Department of Entomology, University of California, Davis 95616, USA.
Hammock
B D
BD
D000818:Animals; D000906:Antibodies / Q000378:metabolism; D015153:Blotting, Western; D004591:Electrophoresis, Polyacrylamide Gel; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004851:Epoxide Hydrolases / Q000037:antagonists & inhibitors / Q000737:chemistry*; D004852:Epoxy Compounds / Q000378:metabolism*; D066298:In Vitro Techniques; D007249:Inflammation / Q000201:enzymology*; D007525:Isoelectric Focusing; D008099:Liver / Q000201:enzymology*; D008264:Macrophages / Q000201:enzymology*; D008297:Male; D051379:Mice; D009504:Neutrophils / Q000201:enzymology*; D011233:Precipitin Tests; D051381:Rats; D011994:Recombinant Proteins / Q000378:metabolism
C096857:1,3-diphenylpropene oxide; D000906:Antibodies; D004791:Enzyme Inhibitors; D004852:Epoxy Compounds; D011994:Recombinant Proteins; D004851:Epoxide Hydrolases