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. 2014 Dec 23;290(8):4928–4940. doi: 10.1074/jbc.M114.601575

FIGURE 2.

FIGURE 2.

Co-receptor binding stabilizes the interaction between PEX7 and its cargo. A, mammalian two-hybrid assay measuring the interaction strength between the PTS2-reporter protein PTS2thiolase-EGFP (VP16AD-PTS2thiolase-EGFP) and either native PEX7 (GAL4DBD-PEX7) or a variant thereof harboring the mutation E287R (GAL4DBD-PEX7E287R). B, modified mammalian two-hybrid assay measuring the interaction strength between GAL4DBD-PTS2thiolase-EGFP and VP16AD-PEX7 or VP16AD-PEX7E287R in the absence or presence of ectopically expressed co-receptor PEX5L. C, modified mammalian two-hybrid assay measuring the interaction strength between VP16AD-PTS2thiolase-EGFP and either GAL4DBD-PEX7 or variants thereof harboring mutations E113R (GAL4DBD-PEX7E113R), E200R (GAL4DBD-PEX7E200R), or E287R (GAL4DBD-PEX7E287R) in the absence or presence of the co-receptor PEX5L. COS7 cells were transfected with expression plasmids for bait and prey proteins, the luciferase and β-galactosidase reporter plasmids, and the expression plasmid encoding PEX5L or the respective empty vector. The ratio of luciferase activity and β-galactosidase activity is indicated.