A new set of assays for the discovery of aminoacyl-tRNA synthetase inhibitors

•Methods to monitor ARS activity: in vitro, in human cells and in bacteria.•Utilization of tRNA suppressors for drug discovery in human cells.•Use of complementation assays to detect species-specific ARS inhibition.•Monitoring ARS activity via ATP consumption and bioluminescence.

Current biochemical methods available to monitor the activity of aminoacyl-tRNA synthetases (ARS) are ill-suited to high-throughput screening approaches for the identification of small-molecule inhibitors of these enzymes. In an attempt to improve the limitations of current assays we have developed a suite of new methods designed to streamline the discovery of new ARS antagonists. This set of assays includes approaches to monitor ARS activity in vitro, in human cells, and in bacteria. They are applicable to several ARSs from any given organism, can be easily adapted to very high-throughput set-ups, and allow for a multi-factorial selection of drug candidates.