Fig 7. Mutation of L143 or C133S does not affect DUB or deISGylase activities. (A) Ubiquitin-TAMRA or ISG15-TAMRA (substrates) were incubated with 10 μ M or 100 nM L pro (wt or indicated mutant), respectively. The change in fluorescent polarization (mFP) was determined over 60 min, with readings recorded every 60 sec. The average trace of assays performed in technical triplicate is shown. Substrate only and KG-TAMRA peptide served as respective negative and positive controls. (B) HEK293T cells were transfected with 0.5 μ g pcDNA-FLAG-RIG-I, 0.5 μ g pcDNA-HA-ubiquitin and 0.1–0.5 μ g pIRES-GFP-L pro . 16 h post transfection, cells were lysed and lysates subjected to Western Blot analysis for FLAG, HA, GFP and