Quantitation of Sirtuin Activity with a One Step Luminescent Assay Poster
Part # PS128
Thomas A. Kirkland1, Nathan J. Evans2 and Andrew L. Niles2
1Promega Biosciences LLC., 277 Granada Dr., San Luis Obispo, CA, 93401
2Promega Corporation, 2800 Woods Hollow Rd., Madison, WI, 53711
AACR 2011 Abstract #4050
The Sirtuin family of histone/protein deacetylatingenzymes has gained considerable interest both for their recognized importance in processes such as gene silencing and expression and in cellular metabolism. We have developed a convenient and sensitive screening methodology to detect and measure both inhibitors and activators of sirtuin activity. Here we describe a one-step, homogeneous luminescent assay for the detection of activity for multiple sirtuin isotypes. The reagent formulation contains a pro-luminescent sirtuin peptide substrate, NAD+, ATP, thermostable luciferase, and a developer enzyme that cleaves deacetylatedlysine residues. The addition of an active sirtuin enzyme source leads to a robust luminescent signal within minutes with “glow-type” characteristics lasting for several hours.
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