a, DSSO crosslinking and mass spectrometry analyses of the UBC4 complex. Each blue line indicates a crosslink between a pair of Lys residues. Note that BBP–Mud2 are crosslinked to Luc7, Prp40, Snu56, and Snu71. b, Co-purification assays probing the interaction between Snu71 (or Prp40) and Luc7. Various combinations of protein A–TEV–Prp40, protein A–TEV–Snu71, and CBP-tagged Luc7 or Luc7ΔCC (with coiled-coil domain (residues 123–190) deleted) were co-overexpressed in yeast (only Snu71 is protein A tagged in the Snu71 + Prp40 lanes), purified using IgG resin, eluted through TEV cleavage, analysed on SDS–PAGE, and visualized using western blot with an anti-CBP antibody to detect Luc7 (top) and Ponceau S stain to show Snu71 or Prp40 (middle). Western blot using the same anti-CBP antibody was used to demonstrate Luc7 expression levels in cell lysates (bottom). The faint band around 26 kD in all lanes of the middle gel is TEV. This experiment was repeated one additional time with similar results. c, The linker (residues 73–131) between the WW and FF domains of Prp40 is predicted to be disordered using program MetaDisorderMD259.