Redesigning the NEDD8 Pathway with a Bacterial Genetic Screen for Ubiquitin-Like Molecule Transfer

Pathways of ubiquitin-like (UBL) molecule transfer regulate a myriad of cellular cascades. Here, we report a high-throughput assay that correlates catalytic human NEDD8 transfer to bacterial survival. The assay was utilized to screen mutant NEDD8 and NAE (NEDD8-activating enzyme) libraries to engineer a more stable NEDD8 and redesign the NEDD8–NAE interface. This approach will be useful in understanding the specificities underlying UBL pathways.