Extended Data Fig. 9: Characterization of the interaction between PEX39 and PEX7, the abundance of PEX39 in yeast, and the interaction of PEX39 with the YG phase.

a, Glutathione-conjugated beads were bound to 1 µM purified recombinant GST-tagged yeast PEX7 or GST alone, and incubated with 1 µM purified recombinant yeast PEX39 fused at the C terminus to GFP (PEX39–GFP) or with GFP alone. Recruitment of each fluorescent protein to the beads was visualized after 1 h on a spinning-disk confocal microscope. Scale bar equals 100 µm. b, Representative Coomassie-stained SDS–PAGE gels corresponding to the binding experiment in Fig. 5d. Briefly, yeast PEX7 was incubated with beads coated with GST-tagged wild-type yeast PEX39 or the indicated mutants; after sedimenting the beads, each supernatant fraction was resolved by SDS–PAGE and stained by Coomassie Blue. Molecular weights (in kD) are marked on the left. c, PEX7 and PEX39 were fused at their C terminus to a FLAG tag (PEX7–FLAG and PEX39–FLAG) and expressed from their respective endogenous promoters in yeast. Shown is a FLAG immunoblot of the total lysate from two different clones (lanes 3 and 4) following growth in oleic acid, compared to cells expressing PEX7–FLAG and untagged PEX39 (lane 2), and cells expressing untagged versions of both proteins (lane 1). Asterisk indicates non-specific bands; molecular weights (in kD) are marked on the left. d, YG hydrogels were incubated with GFP alone or with GFP fused to the C terminus of the indicated proteins. The accumulation of each fluorescent protein inside the gel after 30 min was visualized on a point-scanning confocal microscope. Representative images are shown; the fold enrichment of each protein relative to buffer, across the imaged field, is plotted above (mean ± range of 3 experiments). The width of each image corresponds to 300 µm. Source numerical data and unprocessed blots are provided.

Source data