A cascading activity-based probe sequentially targets E1–E2–E3 ubiquitin enzymes 

Monique P C Mulder, Katharina Witting, Ilana Berlin, Jonathan N Pruneda, Kuen-Phon Wu, Jer-Gung Chang, Remco Merkx, Johanna Bialas, Marcus Groettrup, Alfred C O Vertegaal, Brenda A Schulman, David Komander, Jacques Neefjes, Farid El Oualid* & Huib Ovaa*. Nature Chemical Biology, 16 May 2016, Advance Online Publication, DOI: 10.1038/NCHEMBIO.2084

Triple E probes™ launch

activity-based probes for E1-E2-E3 cascade

We proudly present our new and revolutionary activity-based probes for the E1-E2-E3 enzyme cascade, or Triple E probes. The probes enable you to crystallize, identify or validate dozens of enzymes, or potential drug targets, involved in protein ubiquitination.

Our Triple E probes represent the first tools that allow the monitoring of full E1-E2-E3 cascade activity.

available options

frequently asked questions

Which of the E1-E2-E3 enzymes is targeted by the Triple E probe?
The probe targets active site cysteine based E1-E2-E3 enzymes.

What about the RING class E3 enzymes, which lack a catalytic site cysteine and thus are not directly targeted by the Triple E probe? 
Highly stable E2-Ub(l)-Dha thioether adducts act as stable (competitive) inhibitors of RING type E3 ligases. This allows studies of the RING class E3 enzymes.

Is the probe transferred to substrates of the E1-E2-E3 enzymes?
As far as we know the Triple E Probe is not transferred to substrates.

How specific is the probe labelling? 
The ubiquitin (-like) substrate context of the probe in combination with the required activation by the E1 mediated adenylation step, makes the probe highly specific for E1-E2-E3 enzymes.

Can the probe be used in cell lysates and live cells?
Yes, see our factsheet or the reference [Mulder et al.] as mentioned above.

Is the probe processed in a native way? 
Yes, see above in figure 1, factsheet or reference [Mulder et al.] as mentioned above.