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Journal of Biomolecular Screening
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A Simplified Scintillation Proximity Assay for Fatty Acid Synthase Activity: Development and Comparison with Other FAS Activity Assays

Nathan W. Bays

Department of In Vitro Sciences, Merck Research Laboratories, Boston, Massachusetts, nathan_bays{at}merck.com

Armetta D. Hill

Department of In Vitro Sciences, Merck Research Laboratories, Boston, Massachusetts

Ilona Kariv

Department of In Vitro Sciences, Merck Research Laboratories, Boston, Massachusetts

Fatty acid synthase (FAS), an essential enzyme for de novo lipogenesis, has been implicated in a number of disease states, including obesity, dyslipidemia, and cancer. To identify small-molecule inhibitors of FAS, the authors developed a bead-based scintillation proximity assay (SPA) to detect the fatty acid products of FAS enzymatic activity. This homogeneous SPA assay discriminates between a radiolabeled hydrophilic substrate of FAS (acetyl-coenzyme A) and the labeled lipophilic products of FAS (fatty acids), generating signal only when labeled fatty acids are present. The assay requires a single addition of unmodified polystyrene imaging SPA beads and can be miniaturized to 384- or 1536-well density with appropriate assay statistics for high-throughput screening. High-potency FAS inhibitors were used to compare the sensitivity of the SPA bead assay with previously described assays that measure FAS reaction intermediates (CoA-SH and NADP +). The advantages and disadvantages of these different FAS assays in small-molecule inhibitor discovery are discussed. (Journal of Biomolecular Screening 2009:636-642)

Key Words: fatty acid synthase • FAS • lipogenesis • scintillation proximity assay • SPA

This version was published on July 1, 2009

Journal of Biomolecular Screening, Vol. 14, No. 6, 636-642 (2009)
DOI: 10.1177/1087057109335746
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