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Protein Translocation: the Sec61/Secyeg Translocon

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Protein Translocation: the Sec61/Secyeg Translocon Dispatch R317 4. Read, B.A., Kegel, J., Klute, M.J., Kuo, A., 11. Quince, C., Lanzen, A., Davenport, R.J., and 17. Mangot, J.F., Domaizon, I., Taib, N., Marouni, N., Lefebvre, S.C., Maumus, F., Mayer, C., Turnbaugh, P.J. (2011). Removing noise from Duffaud, E., Bronner, G., and Debroas, D. (2013). Miller, J., Monier, A., Salamov, A., et al. (2013). pyrosequenced amplicons. BMC Short-term dynamics of diversity patterns: Pan genome of the phytoplankton Emiliania Bioinformatics 12, 38. evidence of continual reassembly within underpins its global distribution. Nature 499, 12. Koeppel, A.F., and Wu, M. (2013). Surprisingly lacustrine small eukaryotes. Environ. Microbiol. 209–213. extensive mixed phylogenetic and ecological 15, 1745–1758. 5. Sogin, M.L., Morrison, H.G., Huber, J.A., Mark signals among bacterial Operational 18. Nolte, V., Pandey, R.V., Jost, S., Medinger, R., Welch, D., Huse, S.M., Neal, P.R., Arrieta, J.M., Taxonomic Units. 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USA 107, 1889–1898. 5881–5886. http://dx.doi.org/10.1016/j.cub.2014.03.029 Protein Translocation: The Sec61/ ribosome binds to cytosolic loops of the translocon, whereupon the signal SecYEG Translocon Caught in the Act sequence mediates pore opening and initiates transfer of the growing The Sec61/SecYEG complex mediates both the translocation of newly polypeptide from the ribosome through synthesized proteins across the membrane and the integration of the channel. Hydrophobic segments transmembrane segments into the lipid bilayer. New cryo-electron microscopy trigger lateral opening of the channel studies show ribosome–channel complexes in action and reveal their repertoire and integrate into the membrane as TM of conformational states. segments. Exactly how these steps work mechanistically is not known. The translocon is composed of Martin Spiess released into the lipid bilayer. From subunits SecY, E, and G in bacteria with extensive biochemical analyses and ten, one, and one or two TM domains, Biological membranes separate crystal structures of the closed, idle respectively, corresponding to Sec61a, cellular compartments, generating and translocon, a general picture of these g, and b in eukaryotes [1]. The first preserving concentration gradients dynamic processes has been pieced crystal structure of an idle translocon, and electrical potentials. How are entire together. Two new studies [3,4] now from Methanocaldococcus jannaschii polypeptides transported across or show cryo-electron microscopy (EM) 10 years ago [5], changed the view of inserted into membranes while structures of translocons in action, the translocation pore dramatically. maintaining the barrier? This task is arrested either at the point of signal Rather than an oligomer of several Sec accomplished by a conserved sequence insertion, polypeptide complexes forming a wide water-filled protein-conducting channel — the translocation, or transmembrane channel, it was found to be a compact SecYEG complex at the plasma segment integration, letting us watch helix bundle of a single heterotrimer membrane of prokaryotes, or the Sec61 the translocon at work more directly with the potential to open a narrow pore translocon at the endoplasmic than ever. This work confirms that the (Figure 1A). The ten TM segments of reticulum of eukaryotes [1,2]. picture that emerged from previous SecY form an hourglass shape with an Ribosomes translating secretory or biochemical data is encouragingly empty vestibule on the cytosolic side membrane proteins are targeted to accurate. and a lumenal cavity occupied by a the translocon by signal peptides. As a hydrophobic signal sequence short hydrophobic helix — the Hydrophilic sequences are threaded emerges from the translating ribosome, so-called plug. The two cavities are through a polar channel, while apolar it is bound by the signal recognition separated by a central constriction of transmembrane (TM) segments stop particle (SRP) and targeted to SRP six apolar amino acid side chains. further translocation and are laterally receptors in the membrane. The SecY appears to be composed of two Current Biology Vol 24 No 8 R318 In the new studies, Park et al. [3] and Gogala et al. [4] used cryo-EM and single-particle analysis of defined translocation intermediates to explore and visualize the ribosome-bound translocon in action. They improved on previous cryo-EM structures [18,19] with a number of elegant tricks. Park et al. [3] produced early translocation intermediates in living Escherichia coli cells by inducing expression of a 100-amino acid peptide with an amino-terminal signal sequence and a carboxy-terminal translational stalling sequence. Engineered cysteines were oxidized to form a stabilizing crosslink between the end of the signal sequence and the plug (with the risk of introducing a structural bias), before solubilization and sequential purification for affinity tags in the ribosome and the translocon. Similarly, Gogala et al. [4] translated stalled Figure 1. The translocon in successive functional states. nascent chains with two amino- Schematic representation of conformational states of a ribosome-bound idle translocon (A) or terminal TM segments followed by a translocons engaged with a signal sequence (B), with a translocating chain (C), or with an translocating chain with or without a inserting TM domain (D). The amino- and carboxy-terminal halves of the translocon are shown third TM domain into dog microsomal in blue and red, respectively; the main gate helices TM2 and TM7 as a blue and a red bar; the plug in yellow; and the nascent chains in purple with the signal and TM helices as purple bars. membranes. Upon solubilization, an Side views onto the lateral gate (with part of the ribosomes in gray) are shown in the upper row affinity tag allowed purification of and top views from the cytosol in the lower row. associated ribosome–translocon complexes, and glycosylation sites confirmed the expected membrane- rather symmetrical halves of five TMs Crosslinking the lateral gate shut spanning state of the substrates. connected on one side, where SecE/ abolished secretory protein Resolutions were sufficient to resolve Sec61g also clamps the structure, translocation, whereas crosslinking a-helices of the translocons and extra leaving a single potential lateral outlet across the gate with a spacer of R5A˚ densities of the nascent chains, on the opposite side between TMs 2/3 could still support translocation, allowing molecular dynamics flexible and 7/8. indicating that transport requires some fitting of structural models into the Clearly, the idle translocon expansion of the pore with slight experimental density maps. represents a closed state, and opening opening of the gate [9].
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