HTRF-Based Assay for Microsomal Prostaglandin E 2 Synthase-1 Activity

Abstract

Microsomal prostaglandin E₂ synthase-1 (mPGES-1) catalyzes the formation of prostaglandin E₂ (PGE₂) from the endoperoxide prostaglandin H₂ (PGH₂). Expression of this enzyme is induced during the inflammatory response, and mouse knockout experiments suggest it may be an attractive target for antiarthritic therapies. Assaying the activity of this enzyme in vitro is challenging because of the unstable nature of the PGH₂ substrate. Here, the authors present an mPGES-1 activity assay suitable for characterization of enzyme preparations and for determining the potency of inhibitor compounds. This plate-based competition assay uses homogenous time-resolved fluorescence to measure PGE₂ produced by the enzyme. The assay is insensitive to DMSO concentration up to 10% and does not require extensive washes after the initial enzyme reaction is concluded, making it a simple and convenient way to assess mPGES-1 inhibition. (Journal of Biomolecular Screening 2008:619-625)

Keywords

prostaglandin synthase microsomal prostaglandin E2 synthase-1 homogenous time-resolved fluorescence prostaglandin E2 prostaglandin H2

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