Escherichia coli Cotranslational Targeting and Translocation

Abstract

Targeting of proteins to their proper location inside the cell, in the membrane or in the extracellular space is crucial to the cell. The targeting information is provided in form of a signal sequence by the protein itself. In Escherichia coli, mostly membrane proteins are targeted cotranslationally via the signal recognition particle (SRP) to the membrane. The SRP and its receptor are both guanosine triphosphatases (GTPases) which handover the ribosome‐nascent chain complex to the protein‐conducting channel. Subsequently, the nascent polypeptide is translocated into or across the membrane. To date, a host of biochemical data and a number of structures of important complexes along the pathway are available which shed light on the molecular mechanism of the targeting and translocation process.

Key Concepts:

  • Nascent polypeptides with signal sequences are recognised by the SRP.

  • The hydrophobicity of the signal sequence determines whether a protein is targeted co‐ or posttranslationally.

  • Cotranslational protein targeting is mediated by the SRP and the SRP receptor. They deliver the translating ribosome to the translocation machinery in the membrane.

  • The SecYEG translocon is a channel that allows hydrophilic polypeptides to cross the membrane. In addition, the channel can open laterally and release transmembrane helices into the lipid bilayer.

  • YidC acts as a membrane protein insertase for small membrane proteins in a Sec‐independent manner. In a Sec‐associated form, YidC is suggested to assist in membrane protein folding.

  • YidC, SecD, SecF and YajC can associate with the SecYEG protein‐conducting channel. The resulting complex is called the ‘holotranslocon’, which is suggested to be responsible for membrane protein integration, folding and complex assembly.

Keywords: membrane protein integration; signal recognition particle; FtsY; Sec translocon; YidC integrase; holo‐translocon; SecA ATPase; signal sequence; ribosome; protein export

Figure 1.

Protein export and membrane protein insertion. Exported proteins are targeted posttranslationally via the SecB chaperone and the SecA ATPase to the translocation machinery in the membrane (left). SecA provides the energy for translocation across the membrane. The signal recognition particle (SRP) and its receptor (FtsY) target nascent membrane proteins cotranslationally to the SecYEG protein‐conducting channel (right). Additional proteins (SecD, SecF, YajC and YidC) can associate with SecYEG and assist in the translocation across or into the membrane.

Figure 2.

Cotranslational targeting. (a) Scheme of the domain architecture of E. coliFfh (the SRP protein) and FtsY (SRP receptor). (b) Atomic model of the E. coliSRP in the extended conformation which is observed when the SRP binds the translating ribosome (2J28.pdb) (Halic et al., ). The Ffh N domain is depicted in red, the GTPase domain in orange, the M domain in yellow, the signal sequence in green and the 4.5S RNA in dark red. (c) Co‐crystal structure of the NG domains of Ffh and FtsY from Thermus aquaticus (1OKK.pdb) (Egea et al., ; Focia et al., ). The two GTPase domains form a composite active site. The two GTP analogues are shown as green spheres. The FtsY NG domain is depicted in blue. (d) Model of the SRP and FtsY conformational states during cotranslational targeting (adapted from Zhang et al., ). First, the ribosome nascent chain complex (cargo) is recognised and tightly bound by the SRP. The cryo‐EM structure of the RNC‐SRP (filtered to 15 Å resolution, Halic et al., ) is depicted with the same colour coding as the cartoon drawings of the other complexes. Subsequently, the SRP and FtsY form a GTP‐independent early intermediate. In the presence of GTP, the early intermediate rearranges into the closed conformation. Rearrangements in the catalytic loops of the GTPase domains activate GTP hydrolysis (activated state). This leads to cargo‐release, that is, handover of the RNC to the SecYEG translocon and dissociation of the SRP–FtsY complex.

Figure 3.

The translocation machinery. The crystal structure of the archaeal SecYEβ (top, 1RHZ.pdb) (Van den Berg et al., ) shown from the cytoplasm (a) and as a side view (cut in the middle of the protein‐conducting channel) (b). SecY is depicted in grey; TM2 and TM7 of SecY that line the lateral gate are highlighted in blue and green, respectively. The inactive translocation pore is sealed by a small helix on the periplasmic site (the plug, depicted in red). The ring of hydrophobic amino acids that seals the translocation pore is highlighted in yellow. (c) Topology of the additional translocation factors SecD, SecF, YajC and YidC. The periplasmic domain of E. coliYidC (3BLC.pdb) (Oliver and Paetzel, ) is the only high‐resolution structural information of these proteins currently available.

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References

Arkowitz RA and Wickner W (1994) SecD and SecF are required for the proton electrochemical gradient stimulation of preprotein translocation. EMBO Journal 13(4): 954–963.

Beck K, Eisner G, Trescher D et al. (2001) YidC, an assembly site for polytopic Escherichia coli membrane proteins located in immediate proximity to the SecYE translocon and lipids. EMBO Reports 2(8): 709–714.

van Bloois E, Dekker HL, Froderberg L et al. (2008) Detection of cross‐links between FtsH, YidC, HflK/C suggests a linked role for these proteins in quality control upon insertion of bacterial inner membrane proteins. FEBS Letters 582(10): 1419–1424.

Breyton C, Haase W, Rapoport TA, Kühlbrandt W and Collinson I (2002) Three‐dimensional structure of the bacterial protein‐translocation complex SecYEG. Nature 418(6898): 662–665.

Doudna JA and Batey RT (2004) Structural insights into the signal recognition particle. Annual Review of Biochemistry 73: 539–557.

Duong F and Wickner W (1997) The SecDFyajC domain of preprotein translocase controls preprotein movement by regulating SecA membrane cycling. EMBO Journal 16(16): 4871–4879.

Economou A, Pogliano JA, Beckwith J, Oliver DB and Wickner W (1995) SecA membrane cycling at SecYEG is driven by distinct ATP binding and hydrolysis events and is regulated by SecD and SecF. Cell 83(7): 1171–1181.

Egea PF, Shan SO, Napetschnig J et al. (2004) Substrate twinning activates the signal recognition particle and its receptor. Nature 427(6971): 215–221.

Eisner G, Moser M, Schäfer U, Beck K and Müller M (2006) Alternate recruitment of signal recognition particle and trigger factor to the signal sequence of a growing nascent polypeptide. Journal of Biological Chemistry 281(11): 7172–7179.

Estrozi LF, Boehringer D, Shan S, Ban N and Schaffitzel C (2010) Cryo‐EM Structure of the E. coli translating ribosome in complex with SRP and its receptor. Nature Structural and Molecular Biology. http://dx.doi.org/10.1038/nsmb.1952.

Focia PJ, Shepotinovskaya IV, Seidler JA and Freymann DM (2004) Heterodimeric GTPase core of the SRP targeting complex. Science 303(5656): 373–377.

Gu SQ, Peske F, Wieden HJ, Rodnina MV and Wintermeyer W (2003) The signal recognition particle binds to protein L23 at the peptide exit of the Escherichia coli ribosome. RNA 9(5): 566–573.

Halic M, Becker T, Pool MR et al. (2004) Structure of the signal recognition particle interacting with the elongation‐arrested ribosome. Nature 427(6977): 808–814.

Halic M, Blau M, Becker T et al. (2006) Following the signal sequence from ribosomal tunnel exit to signal recognition particle. Nature 444(7118): 507–511.

Hessa T, Meindl‐Beinker NM, Bernsel A et al. (2007) Molecular code for transmembrane‐helix recognition by the Sec61 translocon. Nature 450(7172): 1026–1030.

Junne T, Schwede T, Goder V and Spiess M (2007) Mutations in the Sec61p channel affecting signal sequence recognition and membrane protein topology. Journal of Biological Chemistry 282(45): 33201–33209.

Kadokura H and Beckwith J (2009) Detecting folding intermediates of a protein as it passes through the bacterial translocation channel. Cell 138(6): 1164–1173.

Keenan RJ, Freymann DM, Stroud RM and Walter P (2001) The signal recognition particle. Annual Review of Biochemistry 70: 755–775.

Kiefer D and Kuhn A (2007) YidC as an essential and multifunctional component in membrane protein assembly. International Review of Cytology 259: 113–138.

Kohler R, Boehringer D, Greber B et al. (2009) YidC and Oxa1 form dimeric insertion pores on the translating ribosome. Molecular Cell 34(3): 344–353.

Lakkaraju AK, Mary C, Scherrer A, Johnson AE and Strub K (2008) SRP keeps polypeptides translocation‐competent by slowing translation to match limiting ER‐targeting sites. Cell 133(3): 440–451.

Lee HC and Bernstein HD (2001) The targeting pathway of Escherichia coli presecretory and integral membrane proteins is specified by the hydrophobicity of the targeting signal. Proceedings of the National Academy of Sciences of the USA 98(6): 3471–3476.

Luirink J, von Heijne G, Houben E and de Gier JW (2005) Biogenesis of inner membrane proteins in Escherichia coli. Annual Review of Microbiology 59: 329–355.

Menetret JF, Schaletzky J, Clemons WM Jr et al. (2007) Ribosome binding of a single copy of the SecY complex: implications for protein translocation. Molecular Cell 28(6): 1083–1092.

Mitra K, Schaffitzel C, Shaikh T et al. (2005) Structure of the E. coli protein‐conducting channel bound to a translating ribosome. Nature 438(7066): 318–324.

Nagai K, Oubridge C, Kuglstatter A et al. (2003) Structure, function and evolution of the signal recognition particle. EMBO Journal 22(14): 3479–3485.

Nagamori S, Smirnova IN and Kaback HR (2004) Role of YidC in folding of polytopic membrane proteins. Journal of Cell Biology 165(1): 53–62.

Neumann‐Haefelin C, Schäfer U, Müller M and Koch HG (2000) SRP‐dependent co‐translational targeting and SecA‐dependent translocation analyzed as individual steps in the export of a bacterial protein. EMBO Journal 19(23): 6419–6426.

Nouwen N and Driessen AJ (2002) SecDFyajC forms a heterotetrameric complex with YidC. Molecular Microbiology 44(5): 1397–1405.

Oliver DC and Paetzel M (2008) Crystal structure of the major periplasmic domain of the bacterial membrane protein assembly facilitator YidC. Journal of Biological Chemistry 283(8): 5208–5216.

Osborne AR and Rapoport TA (2007) Protein translocation is mediated by oligomers of the SecY complex with one SecY copy forming the channel. Cell 129(1): 97–110.

Pogliano JA and Beckwith J (1994) SecD and SecF facilitate protein export in Escherichia coli. EMBO Journal 13(3): 554–561.

Pop OI, Soprova Z, Koningstein G et al. (2009) YidC is required for the assembly of the MscL homopentameric pore. FEBS Journal 276(17): 4891–4899.

Poritz MA, Bernstein HD, Strub K et al. (1990) An E. coli ribonucleoprotein containing 4.5S RNA resembles mammalian signal recognition particle. Science 250(4984): 1111–1117.

Powers T and Walter P (1997) Co‐translational protein targeting catalyzed by the Escherichia coli signal recognition particle and its receptor. EMBO Journal 16(16): 4880–4886.

Rapoport TA (2007) Protein translocation across the eukaryotic endoplasmic reticulum and bacterial plasma membranes. Nature 450(7170): 663–669.

Rehling P, Model K, Brandner K et al. (2003) Protein insertion into the mitochondrial inner membrane by a twin‐pore translocase. Science 299(5613): 1747–1751.

Romisch K, Webb J, Herz J et al. (1989) Homology of 54 K protein of signal‐recognition particle, docking protein and two E. coli proteins with putative GTP‐binding domains. Nature 340(6233): 478–482.

Samuelson JC, Chen M, Jiang F et al. (2000) YidC mediates membrane protein insertion in bacteria. Nature 406(6796): 637–641.

Schaffitzel C, Oswald M, Berger I et al. (2006) Structure of the E. coli signal recognition particle bound to a translating ribosome. Nature 444(7118): 503–506.

Shan SO and Walter P (2005) Co‐translational protein targeting by the signal recognition particle. FEBS Letters 579(4): 921–926.

Simon SM and Blobel G (1992) Signal peptides open protein‐conducting channels in E. coli. Cell 69(4): 677–684.

Skach WR (2009) Cellular mechanisms of membrane protein folding. Nature Structural and Molecular Biology 16(6): 606–612.

Snapp EL, Reinhart GA, Bogert BA, Lippincott‐Schwartz J and Hegde RS (2004) The organization of engaged and quiescent translocons in the endoplasmic reticulum of mammalian cells. Journal of Cell Biology 164(7): 997–1007.

Tam PC, Maillard AP, Chan KK and Duong F (2005) Investigating the SecY plug movement at the SecYEG translocation channel. EMBO Journal 24(19): 3380–3388.

Van den Berg B, Clemons WM Jr, Collinson I et al. (2004) X‐ray structure of a protein‐conducting channel. Nature 427(6969): 36–44.

Walter P and Johnson AE (1994) Signal sequence recognition and protein targeting to the endoplasmic reticulum membrane. Annual Review of Cell Biology 10: 87–119.

Weiche B, Burk J, Angelini S et al. (2008) A cleavable N‐terminal membrane anchor is involved in membrane binding of the Escherichia coli SRP receptor. Journal of Molecular Biology 377(3): 761–773.

Xie K and Dalbey RE (2008) Inserting proteins into the bacterial cytoplasmic membrane using the Sec and YidC translocases. Nature Reviews. Microbioloby 6(3): 234–244.

Zhang X, Schaffitzel C, Ban N and Shan SO (2009) Multiple conformational switches in a GTPase complex control co‐translational protein targeting. Proceedings of the National Academy of Sciences of the USA 106(6): 1754–1759.

Further Reading

Cross BC, Sinning I, Luirink J and High S (2009) Delivering proteins for export from the cytosol. Nature Reviews Molecular Cell Biology 10(4): 255–264.

Facey SJ and Kuhn A (2010) Biogenesis of bacterial inner‐membrane proteins. Cellular and Molecular Life Sciences 67: 2343–2362.

Gasper R, Meyer S, Gotthardt K, Sirajuddin M and Wittinghofer A (2009) It takes two to tango: regulation of G proteins by dimerization. Nature Reviews Molecular Cell Biology 10(6): 423–429.

Rapoport TA (2008) Protein transport across the endoplasmic reticulum membrane. FEBS Journal 275(18): 4471–4478.

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von Loeffelholz, Ottilie, Botte, Matthieu, and Schaffitzel, Christiane(Jan 2011) Escherichia coli Cotranslational Targeting and Translocation. In: eLS. John Wiley & Sons Ltd, Chichester. http://www.els.net [doi: 10.1002/9780470015902.a0023170]