E1 activity

The  E1 LITE - UBE1 Activity Assay Kit is based on the FRET technology.
 
The reaction is made between donor and acceptor fluorophores associated with the two ubiquitin molecules bound to the activated UBE1 E1 LITE - UBE1 Activity Assay Kit from LifeSensorsternary complex (see figure below).
 
 
The assay employs streptavidin terbium as the donor fluorophore, which is directed to the complex by biotinylated ubiquitin and fluoroscein labeled ubiquitin as the acceptor fluorophore. The long-lived fluorescence of the terbium donor allows a time-delayed reading of the FRET signal which reduces the contribution of short-lived background emissions resulting from from buffers, proteins, and chemical compounds.
 
The assay can be run in a 96-well or 384-well format and produces a reproducible signal with a  Z' > 0.7.

Applications

  • Screening for inhibitors of the ubiquitin E1 enzyme, UBE1...
  • Characterization of UBE1 kinetics and of the mechanisms of E1 inhibitors
  • Secondary screening of  E3 ligase, kinase, or other target inhibitors for inhibition of UBE1

Benefits

  • Eliminates the need for radioactive or Western blot-based detection of E1- ubiquitin-thioester formation
  • Homogenous assay compatible with high-throughput screening for UBE1 inhibitors
  • Compatible with 96-well and 384-well formats

Available formats

  • 96 well plate
  • 384 well plate
 
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