We developed bacterial genetic tools to study the Ub system (for full description read the Nature Methods paper).
Ubiquitin (Ub) and ubiquitylation target were fused to split antibiotic resistance fragments via long flexible but proteolytic stable stable linkers. Co-expression of these two chimeras along with functional ubiquitylation apparatus in E. coli give rise to bacterial growth on selective media (supplemented with antibiotic).
We demonstrated the system applications in identifying and characterizing new:
Ubiquitylation targets of given E3
Identification of Sem1 as ubiquitylation target of Rsp5
Quantification of the ubiquitylation dependent bacterial growth