Figure 1.
Additional data supporting our original publication. A, effect of various Tris concentrations on the deglycation assayed as in Ref. 3, measured as the drop in absorption at 290 nm, which detects the hemithioacetal formed by MG reacting with glutathione. B, gel filtration of the HIS-hDJ-1 protein sample used in our paper (2), using a HiPrep 16/60 Sephacryl S-200 HR column, reveals that the protein is not aggregated. Eluates were collected from the column, and the protein concentration in each eluate was measured using the micro BCATM Protein Assay Kit from Thermo Fisher. Aggregated protein would elute with the void volume under 40 ml. Rough estimates for elution volumes of three reference proteins are indicated. C, addition of 1 mm methylglyoxal to S2 cells results in increased levels of the MG adduct CEL only in Glo1 knockdown cells but not in DJ-1β knockdown cells. S2 cells were treated with indicated MG concentrations for 24 h prior to lysis and immunoblotting.