Fluorogenic C-terminal ubiquitin derivatives can be utilised as substrates for deubiquitinating enzymes (DUB), including some members of the ubiquitin specific protease (USP) and ubiquitin C-terminal hydrolase (UCH) enzyme families. Incubation with suitable ubiquitin protease results in release of Rhodamine 110 leading to an increase in fluorescence which can be used in the continuous measurement of DUB activity. Such assays can be used to assess not only the activity of a specific DUB but the suitability of Ub-Rho110 as a substrate and the effect of potential DUB modulators on its activity.
Ub-Rho110 facilitates high sensitivity detection of DUB activity at longer wavelengths than Ub-AMC
Fluorescence measurement: Ex 485nm, Em 535nm wavelengths
