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bioRxiv · 10.1101/148858

Saccharomyces cerevisiae as a platform for assessing sphingolipid kinase inhibitors

Kharel, Y.·Agah, S.·Mendelson, A. J.·Eletu, O. T.·Gesualdi, J.·Smith, J. S.·Santos, W. L.·Lynch, K. R.
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Abstract

Successful medicinal chemistry campaigns to discover and optimize sphingosine kinase inhibitors require a robust assay for screening chemical libraries and for determining rank order potencies. Existing assays for these enzymes are laborious, expensive and/or low throughput. The toxicity of excessive levels of phosphorylated sphingoid bases for the budding yeast, Saccharomyces cerevisiae, affords an assay wherein inhibitors added to the culture media rescue growth in a dose-dependent fashion. Herein, we describe our adaptation of a simple, inexpensive, and high throughput assay for assessing inhibitors of sphingosine kinase types 1 and 2 as well as ceramide kinase and for testing enzymatic activity of sphingosine kinase type 2 mutants. The assay was validated using recombinant enzymes and generally agrees with rank order of potencies of existing inhibitors.

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Kharel, Y., Agah, S., Mendelson, A. J., Eletu, O. T., Gesualdi, J., Smith, J. S., Santos, W. L., Lynch, K. R.. 2017-06-11. Saccharomyces cerevisiae as a platform for assessing sphingolipid kinase inhibitors. https://doi.org/10.1101/148858

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