Fig. 4. The C-terminal tail of Sys1p (amino acids 151–203) interacts with the Sec23p–Sec24p complex. GST and a GST–Sys1 tail fusion protein, coupled to agarose beads, served as affinity matrix with detergent-solubilized total yeast protein. Beads were washed with buffer, and bound proteins eluted in 1 ml fractions of SDS buffer (1) or 0.5 ml fractions of 1 M NaCl (2) for SDS–PAGE and silver staining (20 µl of each fraction). Bound proteins usually appeared in two fractions, one of which is shown on the blot. Asterisks mark two unspecifically bound proteins of ∼40 and 50 kDa.