TitleAuthorsAbstractStructure of the translocation channelDifferent modes of translocationOpening the channel across the membraneThe poreOligomeric translocation channelsMembrane-protein integrationMaintaining the permeability barrierPerspectiveReferencesFigure 1 The translocation channel.Figure 2 Model of co-translational translocation.Figure 3 Model of post-translational translocation in eukaryotes.Figure 4 Model of post-translational translocation in bacteria.Figure 5 Different stages of translocation.REVIEWSProtein translocation across the eukaryoticendoplasmic reticulum and bacterialplasma membranesTom A. Rapoport1A decisive step in the biosynthesis of many proteins is their partial or complete translocation across the eukaryoticendoplasmic reticulum membrane or the prokaryotic plasma membrane. Most of these proteins are transloca ted through aprotein-conducting channel that is formed by a conserved, heterotrimeric membrane-protein complex, the Sec61 or SecYcomplex. Depending on channel binding partners, polypeptides are moved by different mechanisms: the polypeptide chain istransferred directly into the channel by the translating ribosome, a ratcheting mechanism is used by the endo plasmicreticulum chaperone BiP, and a pushing mechanism is used by the bacterial AT Pase SecA. Str uctural, genetic andbiochemical data show how the channel opens across the membrane, releases hydrophobic segments of membrane proteinslaterally into lipid, and maintains the membrane barrier for small mole cules.For almost 40 years, researchers have been fascinated by thequestion of how proteins are transported across or are inte-grated into membranes. Pioneering work by G. Palade1demonstrated that in eukaryotic cells secretory proteins crossthe endoplasmic reticulum membrane before being transported invesicles to the plasma membrane. The laboratories of G. Blobel and C.Milstein then discovered that these proteins are directed to the endo-plasmic reticulum membrane by signal sequences2,3. A little later,signal sequences were also found to direct the translocation of pro-teins across the bacterial plasma membrane4,5. Genetic experimentsidentified components required for translocation, initially in bacteriaand later in yeast6–8, and the establishment of an in vitro systeminitiated biochemical studies9. All of these achievements set the stagefor investigations into the molecular mechanism of translocation,which will be the focus of this review.Proteins transported across the eukaryotic endoplasmic reticulummembrane or the prokaryotic plasma membrane include solubleproteins, such as those ultimately secreted from the cell or localizedto the endoplasmic reticulum lumen, and membrane proteins, suchas those in the plasma membrane or in other organelles of the secret-ory pathway. Soluble proteins cross the membrane completely andusually have amino-terminal, cleavable signal sequences, the majorfeature of which is a segment of 7–12 hydrophobic amino acids.Membrane proteins have different topologies in the lipid bilayer,with one or more transmembrane segments composed of about20 hydrophobic amino acids; the hydrophilic regions of these pro-teins either cross the membrane or remain in the cytosol. Both typesof proteins are handled by the same machinery within the membrane:a protein-conducting channel. The channel allows soluble polypep-tides to cross the membrane and hydrophobic transmembrane seg-ments of membrane proteins to exit laterally into the lipid phase.Structure of the translocation channelThe translocation channel is formed from a conserved hetero-trimeric membrane protein complex, called the Sec61 complex ineukaryotes and the SecY complex in bacteria and archaea (for moredetails, see refs 10 and 11). The a- and c-subunits show significantsequence conservation, and both subunits are essential for the func-tion of the channel and for cell viability. The b-subunits are notessential; they are similar in eukaryotes and archaea, but show noobvious homology to the corresponding subunit in bacteria.The a-subunit forms the pore of the channel, as initially shown byexperiments in which photoreactive probes were systematicallyplaced at different positions of a stalled translocating polypeptide12;all positions predicted to be within the membrane cross-linked onlyto the a-subunit of the Sec61 complex, indicating that this subunitsurrounds the polypeptide chain during its passage across the mem-brane. In addition, experiments in which the purified Sec61/SecYcomplex was reconstituted into proteoliposomes showed that it isthe essential membrane component for protein translocation13–15.The channel has an aqueous interior, as demonstrated by electro-physiology experiments16and by measurements of the fluorescencelifetime of probes incorporated into a translocating polypeptidechain17,18.The crystal structure of an archaeal SecY complex providedimportant insight into how the a-subunit forms the channel10. Thestructure is probably representative of complexes from all species, asindicated by sequence conservation and by the similarity to a lower-resolution structure of the Escherichia coli SecY complex, determinedby electron microscopy from two-dimensional crystals19,20. Viewedfrom the cytosol, the channel has a square shape (Fig. 1a). Thea-subunit is divided into two halves, transmembrane segments 1–5and 6–10. The loop between transmembrane segments 5 and 6 at theback of the a-subunit serves as a hinge, allowing the a-subunit toopen at the front—the ‘lateral gate’. The c-subunit links the twohalves of the a-subunit at the back by extending one transmembranesegment diagonally across their interface. The b-subunit makes con-tact only with the periphery of the a-subunit, probably explainingwhy it is dispensable for the function of the complex.The ten helices of the a-subunit form an hourglass-shaped porethat consists of cytoplasmic and external funnels, the tips of whichmeet about half way across the membrane (Fig. 1b). Whereas the1Howard Hughes Medical Institute and Department of Cell Biology, Harvard Medical School, 240 Longwood Avenue, Boston, Massachusetts 02115, USA.Vol 450j29 November 2007jdoi:10.1038/nature06384663Nature ©2007PublishingGroupcytoplasmic funnel is empty, the external funnel is plugged by a shorthelix. The crystal structure therefore represents a closed channel but,as will be discussed later, biochemical data indicate how it can openand translocate proteins. The constriction of the