Fig. 4

#56 analogs target SOD1 DBR. a In vitro IP assay with various SOD1-Derlin-1 complexes. SOD1-Derlin-1 complexes purified from lysates transfected with the indicated plasmids were incubated with #56-20 or #56-26 for 16 h and analyzed by IP-IB with the indicated antibodies. b FP analysis of #56-20 in the presence of indicated concentration of SOD1 (5-20) and Derlin-1 (CT4) peptide. Here, 250 nM #56-20 was incubated with the indicated concentration of each peptide for 6 h, and FP was measured. Blue: Derlin-1 (CT4); red: SOD1 (5-20). The data are shown as the mean ± s.d. (n = 3). c–e Pull-down assay using PB56. HEK293A cells transfected with indicated plasmid were treated with 200 μM PB56 for 24 h followed by UV light irradiation for 1 h. Lysates were analyzed by IP-IB with the indicated antibodies. c Pull-down assay by Flag-SOD1^G93A or Derlin-1-Flag. Chemical structure shows PB56 (blue: #56-59 (active part); black: benzophenone with polyethylene glycol (PEG) as spacers; red: biotin). d Competition assay with the pre-treatment of 20 μM #56-59 for 12 h. e Pull-down assay by Flag-SOD1^G93A or Flag-SOD1^WT