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Detection of JumonjiC Domain-Containing Histone Demethylase Activities with Homogeneous Bioluminescent Succinate Assay Poster

Part # PS246

Abstract

Richard Somberg, Juliano Alves, Said Goueli, and Hicham Zegzouti
Promega Corporation, 2800 Woods Hollow Rd, Madison, WI, 53711

JumonjiC domain-containing histone lysine demethylases (JMJCs) play a pivotal role in determining the epigenetic status of the genome, and because of their implication in cancer they have become validated drug targets. Since the conversion of 2-oxoglutarate to succinate is a required step for the activity of these enzymes, an assay that detects succinate would be suitable for monitoring all JMJCs. We therefore developed a homogeneous bioluminescent detection assay that is performed in a two-step format, and relies on converting succinate to ATP and light in a robust luciferase reaction. The assay is highly sensitive and robust, the light output is proportional to succinate concentration, and the method was validated with several JMJC enzymes and inhibitors. Therefore, the succinate detection assay is a simple-to-use method that allows significant savings of enzyme usage and does not require antibodies or modified substrates, and could have significant impact on diverse areas of epigenetics research.

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