Figure 6.

The Sec complex is intact in Cs sec61 strains. ER membrane proteins were fractionated after solubilization in digitonin. Total, total microsomal protein; Sol., digitonin-soluble protein recovered from the 100,000 × g supernatant; Free, fraction of digitonin-soluble proteins not binding to Con-A; Con-A, proteins binding to concanavalin A. Equal aliquots of each fraction were analyzed by SDS-PAGE and immunoblotting using the indicated antibodies in the first incubation. For Sec63p and Sec61p, [³⁵S]protein A was used in the secondary incubation, and blots were exposed to a PhosphorImager. Sec72p was visualized using the ECL procedure. The band visible below Sec63p in the Sec63p immunoblots is unrelated.