Figure 6.

UbFluor can quantify PARKIN activation or inactivation by known point mutations. A, UbMES assay on ΔUbl, ΔUPD, and ΔUPD (C431A) and activating PARKIN mutations in the ΔUbl background (A398T, W403A, and F463Y). Experiments with each mutant were performed using 30 μm UbMES and 1.0 μm PARKIN for the time indicated. W403A and F463Y were run on a separate gel. B, bar graph showing bimolecular reaction rates of UbFluor consumption under MT conditions by PARKIN mutants as described above using the UbFluor assay. Errors are based on linear fits of data from two repeats of UbFluor consumption data at four different concentrations of UbFluor (eight total measurements).