Many proteins synthesized in the cytoplasm of a cell are secreted outside the cell or are transported into specific cellular compartments. For proteins that are secreted or targeted to the membrane, the sorting information is usually encoded in the N-terminal segment of the protein, called the signal sequence. Signal sequences vary in length and actual amino acid sequence, but share a common feature — a central hydrophobic patch flanked on either side by polar regions. Interestingly, in prokaryotes, secreted proteins and membrane proteins containing multiple transmembrane helices (polytopic membrane proteins) appear to be translocated via different pathways — targeting of secreted proteins requires the proteins SecA and SecB, while integration of polytopic membrane proteins involves an RNA–protein complex, called the signal sequence recognition particle (SRP). Thus, one intriguing question is: how does the translocation machinery interpret the signal sequences and thereby target the proteins to the correct locations?

To address this question, Beck et al. (EMBO J., 19: 134–143) used an in vitro translation system and crosslinking techniques to identify ribosome-associated factors that can distinguish between a polytopic membrane protein and a secreted protein. Their results indicate that the translocation pathway is selected early during protein synthesis, with two factors playing a determining role. For a polytopic membrane protein (left panel), the signal sequence (blue box) emerging from the ribosome (brown ellipsoids) binds to SRP (green ellipsoid) and the subsequent membrane integration occurs simultaneously with protein synthesis. In contrast, trigger factor (Tig, red ellipsoid), a chaperone protein tightly associated with the ribosome, binds to the nascent chain of a secreted protein (right panel) and prevents SRP binding to the signal sequence (blue hashed box). As a result, the protein is directed to the SecA/SecB-dependent translocation pathway after synthesis is complete.

The results of Beck et al. suggest that trigger factor and SRP bind to different regions of the nascent polypeptide chain. However, despite having different interaction sites, binding of these two factors appears mutually exclusive. Their study therefore leads to new questions. What are the features recognized by trigger factor, and is this recognition sequence-specific? Does trigger factor binding persist throughout protein synthesis? How does trigger factor binding exclude SRP–signal sequence interactions? A better understanding of how proteins are targeted to different compartments will emerge as these questions are addressed.