Thousands of cellular reactions use nucleotides, including most of the enzymes that catalyze posttranslational modifications of proteins such as phosphorylation and methylation. Because of their central role in signal transduction, many PTM enzymes have become drug targets. Though protein kinases get the most attention, acetyltransferases, methyltransferases, and glycosyltransferases are also clinically validated targets, and many more PTM-related drugs are in the pipeline. Transcreener assays were developed to enable HTS efforts targeting PTM enzymes, as well as other types of nucleotide-dependent enzymes, including ATPases, GTPases, and phosphodiesterases.
Transcreener Assays were developed and continuously improved for use in an industrial HTS setting. Since their introduction in 2006, they have been used worldwide in over 60M wells of inhibitor screening, profiling and mechanistic studies in 96, 384 or 1536 well plates with all of the major multimode plate readers. Use of ADP detection for kinase assays accounts for more than half of those wells, and most of these users decided on Transcreener after comparing it to alternative assay methods. For many more atypical targets – glycosyltransferases, helicases, carboxylases, GTPases, ligases, to name a few – there are no good alternatives, and Transcreener has been a truly enabling technology. The screen would likely not have gone forward without it.
Regardless of the target, all of the Transcreener assays generate Z’ values of greater than 0.7 at substrate conversion levels of less than 10% – usually far lower – over the full range of initial substrate concentration. They are largely unaffected by most reagents used in enzyme assays, and the signals are rock solid at least overnight. We have collaborated with the major multimode instrument providers including Tecan, BMG, Biotek and Molecular Devices to optimize instrument hardware and software settings for maximal performance with each of the Transcreener Assays. Optimal filter sets and instrument settings are summarized in the Technical Manual for each Transcreener assay, and more detailed information is available in Application Notes. This ensures that whatever fluorescent detection mode or reader you prefer, you will get robust results with Transcreener assay reagents.