Development of fluorescent substrates for microsomal epoxide hydrolase and application to inhibition studies

Christophe Morisseau, Maud Bernay, Aurélie Escaich, James R. Sanborn, Jozsef Lango, Bruce D. Hammock

Research output: Contribution to journalArticle

14 Scopus citations

Abstract

The microsomal epoxide hydrolase (mEH) plays a significant role in the metabolism of numerous xenobiotics. In addition, it has a potential role in sexual development and bile acid transport, and it is associated with a number of diseases such as emphysema, spontaneous abortion, eclampsia, and several forms of cancer. Toward developing chemical tools to study the biological role of mEH, we designed and synthesized a series of absorbent and fluorescent substrates. The highest activity for both rat and human mEH was obtained with the fluorescent substrate cyano(6-methoxy-naphthalen-2-yl)methyl glycidyl carbonate (11). An in vitro inhibition assay using this substrate ranked a series of known inhibitors similarly to the assay that used radioactive cis-stilbene oxide but with a greater discrimination between inhibitors. These results demonstrate that the new fluorescence-based assay is a useful tool for the discovery of structure-activity relationships among mEH inhibitors. Furthermore, this substrate could also be used for the screening chemical library with high accuracy and with a Z′ value of approximately 0.7. This new assay permits a significant decrease in labor and cost and also offers the advantage of a continuous readout. However, it should not be used with crude enzyme preparations due to interfering reactions.

Original languageEnglish (US)
Pages (from-to)154-162
Number of pages9
JournalAnalytical Biochemistry
Volume414
Issue number1
DOIs
StatePublished - Jul 1 2011

Keywords

  • α-Cyanocarbonate
  • Fluorescent substrate
  • Microsomal epoxide hydrolase
  • Xenobiotic metabolism

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Molecular Biology
  • Cell Biology

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