| Abstract: |
The Sec61p complex forms the core element of the protein translocation complex (translocon) in the rough endoplasmic reticulum (rough ER) membrane. Translating or nontranslating ribosomes bind with high affinity to ER membranes that have been stripped of ribosomes or to liposomes containing purified Sec61p. Here we present evidence that the β subunit of the complex (Sec61β) makes contact with nontranslating ribosomes. A fusion protein containing the Sec61β cytoplasmic domain (Sec61βc) prevents the binding of ribosomes to stripped ER-derived membranes and also binds to ribosomes directly with an affinity close to the affinity of ribosomes for stripped ER-derived membranes. The ribosome binding activity of Sec61βc, like that of native ER membranes, is sensitive to high salt concentrations and is not based on an unspecific charge-dependent interaction of the relatively basic Sec61βc domain with ribosomal RNA. Like stripped ER membranes, the Sec61βc sequence binds to large ribosomal subunits in preference over small subunits. Previous studies have shown that Sec61β is inessential for ribosome binding and protein translocation, but translocation is impaired by the absence of Sec61β, and it has been proposed that Sec61β assists in the insertion of nascent proteins into the translocation pore. Our results suggest a physical interaction of the ribosome itself with Sec61β; this may normally occur alongside interactions between the ribosome and other elements of Sec61p, or it may represent one stage in a temporal sequence of binding. |
