Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Review Article
  • Published:

Lipopolysaccharide transport and assembly at the outer membrane: the PEZ model

Nature Reviews Microbiology volume 14, pages 337–345 (2016)Cite this article

Subjects

Key Points

  • The outer membrane (OM) of most Gram-negative bacteria contains lipopolysaccharide (LPS), a large molecule that contains several fatty acyl chains and up to hundreds of sugars, in its outer leaflet, which creates a barrier that prevents the entry of large polar molecules and small hydrophobic molecules.

  • The transport of millions of LPS molecules from the inner membrane (IM), across the aqueous periplasmic compartment, and across the OM to the cell surface was not well understood, except that the process is mediated by seven essential and conserved LPS transport (Lpt) proteins.

  • The extraction of LPS from the IM is mediated by an ATP binding cassette (ABC) transporter, LptB2FG, and an associated membrane protein, LptC. These proteins couple ATP hydrolysis in the cytoplasm by LptB to movement to LptC; the LptB2FG and LptB2FGC protein complexes have been purified and demonstrate ATPase activity in vitro.

  • LPS is thought to transit the periplasm by a bridge between LptC and the OM mediated by the periplasmic protein LptA. The bridge is formed by structurally homologous domains of LptC, LptA and the OM protein LptD, and it helps to mediate the transit of the hydrophobic acyl chains of LPS through an aqueous compartment.

  • The OM β-barrel protein LptD and the OM lipoprotein LptE form a two-protein plug-and-barrel complex that is responsible for transporting LPS from the periplasmic bridge across the OM to the cell surface. A current model proposes that the OM translocon changes its conformation, enabling LPS molecules to enter the barrel of LptD and move to the cell surface through lateral openings without ever residing in the inner leaflet of the OM.

  • LptD is a large β-barrel protein that contains two non-consecutive disulfide bonds, either of which is sufficient for the function of LptD. Correct rearrangement of the disulfide bonds to the final configuration is required for LptA to interact with LptD, which prevents mislocalization of LPS when the OM translocon is not properly assembled.

  • Identification of LPS transport intermediates in Escherichia coli cells has enabled the development of a system to study the ATP requirement for LPS transport out of membrane vesicles to soluble LptA. Using this system, the PEZ model was developed to describe how ATP hydrolysis by LptB in the cytoplasm 'pushes' LPS molecules in a continuous stream out of the IM toward the cell surface through the periplasmic bridge comprised of LptC, LptA and LptD.

Abstract

Gram-negative bacteria have a double-membrane cellular envelope that enables them to colonize harsh environments and prevents the entry of many clinically available antibiotics. A main component of most outer membranes is lipopolysaccharide (LPS), a glycolipid containing several fatty acyl chains and up to hundreds of sugars that is synthesized in the cytoplasm. In the past two decades, the proteins that are responsible for transporting LPS across the cellular envelope and assembling it at the cell surface in Escherichia coli have been identified, but it remains unclear how they function. In this Review, we discuss recent advances in this area and present a model that explains how energy from the cytoplasm is used to power LPS transport across the cellular envelope to the cell surface.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: LPS transport pathway in Escherichia coli.
Figure 2: The periplasmic protein bridge.
Figure 3: The outer membrane translocon.
Figure 4: Regulation of the formation of the Lpt bridge.
Figure 5: The PEZ model.

Similar content being viewed by others

Accession codes

Accessions

Protein Data Bank

References

  1. Nikaido, H. Molecular basis of bacterial outer membrane permeability revisited. Microbiol. Mol. Biol. Rev. 67, 593–656 (2003).

    CAS  PubMed  PubMed Central  Google Scholar 

  2. Berg, H. C. The rotary motor of bacterial flagella. Annu. Rev. Biochem. 72, 19–54 (2003).

    CAS  PubMed  Google Scholar 

  3. Nan, B., McBride, M. J., Chen, J., Zusman, D. R. & Oster, G. Bacteria that glide with helical tracks. Curr. Biol. 24, R169–R173 (2014).

    CAS  PubMed  PubMed Central  Google Scholar 

  4. Lai, Y., Rosenshine, I., Leong, J. M. & Frankel, G. Intimate host attachment: enteropathogenic and enterohaemorrhagic Escherichia coli. Cell. Microbiol. 15, 1796–1808 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  5. Laverty, G., Gorman, S. P. & Gilmore, B. F. Biomolecular mechanisms of Pseudomonas aeruginosa and Escherichia coli biofilm formation. Pathogens 3, 596–632 (2014).

    CAS  PubMed  PubMed Central  Google Scholar 

  6. Raetz, C. R. & Whitfield, C. Lipopolysaccharide endotoxins. Annu. Rev. Biochem. 71, 635–700 (2002).

    CAS  PubMed  Google Scholar 

  7. Garten, W., Hindennach, I. & Henning, U. The major proteins of the Escherichia coli outer cell-envelope membrane. Cyanogen bromide fragments of protein I, composition and order. Eur. J. Biochem. 60, 303–307 (1975).

    CAS  PubMed  Google Scholar 

  8. Kamio, Y. & Nikaido, H. Outer membrane of Salmonella typhimurium: accessibility of phospholipid head groups to phospholipase c and cyanogen bromide activated dextran in the external medium. Biochemistry 15, 2561–2570 (1976). This study is the first to demonstrate that phospholipids are not exposed on the cell surface of Gram-negative bacteria.

    CAS  PubMed  Google Scholar 

  9. Osborn, M. J., Gander, J. E., Parisi, E. & Carson, J. Mechanism of assembly of the outer membrane of Salmonella typhimurium. Isolation and characterization of cytoplasmic and outer membrane. J. Biol. Chem. 247, 3962–3972 (1972). This study demonstrates that LPS fractionates almost exclusively with the OM following sucrose-density gradient centrifugation.

    CAS  PubMed  Google Scholar 

  10. Sutcliffe, I. C. A phylum level perspective on bacterial cell envelope architecture. Trends Microbiol. 18, 464–470 (2010).

    CAS  PubMed  Google Scholar 

  11. Ruiz, N., Kahne, D. & Silhavy, T. J. Transport of lipopolysaccharide across the cell envelope: the long road of discovery. Nat. Rev. Microbiol. 7, 677–683 (2009).

    CAS  PubMed  PubMed Central  Google Scholar 

  12. Raetz, C. R., Reynolds, C. M., Trent, M. S. & Bishop, R. E. Lipid A modification systems in Gram-negative bacteria. Annu. Rev. Biochem. 76, 295–329 (2007).

    CAS  PubMed  PubMed Central  Google Scholar 

  13. Whitfield, C. & Trent, M. S. Biosynthesis and export of bacterial lipopolysaccharides. Annu. Rev. Biochem. 83, 99–128 (2014).

    CAS  PubMed  Google Scholar 

  14. Osborn, M. J., Gander, J. E. & Parisi, E. Mechanism of assembly of the outer membrane of Salmonella typhimurium. Site of synthesis of lipopolysaccharide. J. Biol. Chem. 247, 3973–3986 (1972). This paper establishes that LPS is synthesized inside the cell, raising the question of how LPS is transported to its final destination.

    CAS  PubMed  Google Scholar 

  15. Karow, M. & Georgopoulos, C. The essential Escherichia coli msbA gene, a multicopy suppressor of null mutations in the htrB gene, is related to the universally conserved family of ATP-dependent translocators. Mol. Microbiol. 7, 69–79 (1993).

    CAS  PubMed  Google Scholar 

  16. Polissi, A. & Georgopoulos, C. Mutational analysis and properties of the msbA gene of Escherichia coli, coding for an essential ABC family transporter. Mol. Microbiol. 20, 1221–1233 (1996).

    CAS  PubMed  Google Scholar 

  17. Zhou, Z., White, K. A., Polissi, A., Georgopoulos, C. & Raetz, C. R. Function of Escherichia coli MsbA, an essential ABC family transporter, in lipid A and phospholipid biosynthesis. J. Biol. Chem. 273, 12466–12475 (1998).

    CAS  PubMed  Google Scholar 

  18. Doerrler, W. T., Gibbons, H. S. & Raetz, C. R. MsbA-dependent translocation of lipids across the inner membrane of Escherichia coli. J. Biol. Chem. 279, 45102–45109 (2004).

    CAS  PubMed  Google Scholar 

  19. Ward, A., Reyes, C. L., Yu, J., Roth, C. B. & Chang, G. Flexibility in the ABC transporter MsbA: Alternating access with a twist. Proc. Natl Acad. Sci. USA 104, 19005–19010 (2007).

    CAS  PubMed  Google Scholar 

  20. McGrath, B. C. & Osborn, M. J. Localization of terminal steps of O-antigen synthesis in Salmonella typhimurium. J. Bacteriol. 173, 649–654 (1991).

    CAS  PubMed  PubMed Central  Google Scholar 

  21. Whitfield, C. Biosynthesis and assembly of capsular polysaccharides in Escherichia coli. Annu. Rev. Biochem. 75, 39–68 (2006).

    CAS  PubMed  Google Scholar 

  22. Cuthbertson, L., Kos, V. & Whitfield, C. ABC transporters involved in export of cell surface glycoconjugates. Microbiol. Mol. Biol. Rev. 74, 341–362 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  23. Greenfield, L. K. & Whitfield, C. Synthesis of lipopolysaccharide O-antigens by ABC transporter-dependent pathways. Carbohydr. Res. 356, 12–24 (2012).

    CAS  PubMed  Google Scholar 

  24. Sperandeo, P., Deho, G. & Polissi, A. The lipopolysaccharide transport system of Gram-negative bacteria. Biochim. Biophys. Acta 1791, 594–602 (2009).

    CAS  PubMed  Google Scholar 

  25. Simpson, B. W., May, J. M., Sherman, D. J., Kahne, D. & Ruiz, N. Lipopolysaccharide transport to the cell surface: biosynthesis and extraction from the inner membrane. Philos. Trans. R. Soc. Lond. B Biol. Sci. 370, 20150029 (2015).

    PubMed  PubMed Central  Google Scholar 

  26. May, J. M., Sherman, D. J., Simpson, B. W., Ruiz, N. & Kahne, D. Lipopolysaccharide transport to the cell surface: periplasmic transport and assembly into the outer membrane. Philos. Trans. R. Soc. Lond. B Biol. Sci. 370, 20150027 (2015).

    PubMed  PubMed Central  Google Scholar 

  27. Sperandeo, P. et al. Functional analysis of the protein machinery required for transport of lipopolysaccharide to the outer membrane of Escherichia coli. J. Bacteriol. 190, 4460–4469 (2008).

    CAS  PubMed  PubMed Central  Google Scholar 

  28. Ruiz, N., Gronenberg, L. S., Kahne, D. & Silhavy, T. J. Identification of two inner-membrane proteins required for the transport of lipopolysaccharide to the outer membrane of Escherichia coli. Proc. Natl Acad. Sci. USA 105, 5537–5542 (2008).

    CAS  PubMed  Google Scholar 

  29. Rietschel, E. T. et al. Bacterial endotoxin: molecular relationships of structure to activity and function. FASEB J. 8, 217–225 (1994).

    CAS  PubMed  Google Scholar 

  30. Okuda, S., Freinkman, E. & Kahne, D. Cytoplasmic ATP hydrolysis powers transport of lipopolysaccharide across the periplasm in E. coli. Science 338, 1214–1217 (2012). This study describes the first observation of intermediate LPS transport states in living cells.

    CAS  PubMed  PubMed Central  Google Scholar 

  31. Sherman, D. J. et al. Decoupling catalytic activity from biological function of the ATPase that powers lipopolysaccharide transport. Proc. Natl Acad. Sci. USA 111, 4982–4987 (2014). This paper provides the first biochemical and genetic evidence that LPS transport is powered by ATP hydrolysis by LptB.

    CAS  PubMed  Google Scholar 

  32. Sperandeo, P. et al. Characterization of lptA and lptB, two essential genes implicated in lipopolysaccharide transport to the outer membrane of Escherichia coli. J. Bacteriol. 189, 244–253 (2007).

    CAS  PubMed  Google Scholar 

  33. Narita, S. & Tokuda, H. Biochemical characterization of an ABC transporter LptBFGC complex required for the outer membrane sorting of lipopolysaccharides. FEBS Lett. 583, 2160–2164 (2009). This study describes the purification of the LptBFGC complex at the IM.

    CAS  PubMed  Google Scholar 

  34. Sherman, D. J., Okuda, S., Denny, W. A. & Kahne, D. Validation of inhibitors of an ABC transporter required to transport lipopolysaccharide to the cell surface in Escherichia coli. Bioorg. Med. Chem. 21, 4846–4851 (2013).

    CAS  PubMed  Google Scholar 

  35. Villa, R. et al. The Escherichia coli Lpt transenvelope protein complex for lipopolysaccharide export is assembled via conserved structurally homologous domains. J. Bacteriol. 195, 1100–1108 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  36. Suits, M. D., Sperandeo, P., Deho, G., Polissi, A. & Jia, Z. Novel structure of the conserved Gram-negative lipopolysaccharide transport protein A and mutagenesis analysis. J. Mol. Biol. 380, 476–488 (2008).

    CAS  PubMed  Google Scholar 

  37. Matsuyama, S., Tajima, T. & Tokuda, H. A novel periplasmic carrier protein involved in the sorting and transport of Escherichia coli lipoproteins destined for the outer membrane. EMBO J. 14, 3365–3372 (1995).

    CAS  PubMed  PubMed Central  Google Scholar 

  38. Okuda, S. & Tokuda, H. Lipoprotein sorting in bacteria. Annu. Rev. Microbiol. 65, 239–259 (2011).

    CAS  PubMed  Google Scholar 

  39. Takeda, K. et al. Crystal structures of bacterial lipoprotein localization factors, LolA and LolB. EMBO J. 22, 3199–3209 (2003).

    CAS  PubMed  PubMed Central  Google Scholar 

  40. Okuda, S., Watanabe, S. & Tokuda, H. A short helix in the C-terminal region of LolA is important for the specific membrane localization of lipoproteins. FEBS Lett. 582, 2247–2251 (2008).

    CAS  PubMed  Google Scholar 

  41. Tefsen, B., Geurtsen, J., Beckers, F., Tommassen, J. & de Cock, H. Lipopolysaccharide transport to the bacterial outer membrane in spheroplasts. J. Biol. Chem. 280, 4504–4509 (2005). This study provides the first evidence that LPS transport does not involve a soluble periplasmic chaperone.

    CAS  PubMed  Google Scholar 

  42. Chng, S. S., Gronenberg, L. S. & Kahne, D. Proteins required for lipopolysaccharide assembly in Escherichia coli form a transenvelope complex. Biochemistry 49, 4565–4567 (2010). This paper provides evidence that LPS transport involves a trans-envelope bridge.

    CAS  PubMed  PubMed Central  Google Scholar 

  43. Ishidate, K. et al. Isolation of differentiated membrane domains from Escherichia coli and Salmonella typhimurium, including a fraction containing attachment sites between the inner and outer membranes and the murein skeleton of the cell envelope. J. Biol. Chem. 261, 428–443 (1986).

    CAS  PubMed  Google Scholar 

  44. Bos, M. P., Robert, V. & Tommassen, J. Biogenesis of the Gram-negative bacterial outer membrane. Annu. Rev. Microbiol. 61, 191–214 (2007).

    CAS  PubMed  Google Scholar 

  45. Freinkman, E., Okuda, S., Ruiz, N. & Kahne, D. Regulated assembly of the transenvelope protein complex required for lipopolysaccharide export. Biochemistry 51, 4800–4806 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  46. Chin, J. W., Martin, A. B., King, D. S., Wang, L. & Schultz, P. G. Addition of a photocrosslinking amino acid to the genetic code of Escherichia coli. Proc. Natl Acad. Sci. USA 99, 11020–11024 (2002).

    CAS  PubMed  Google Scholar 

  47. Liu, C. C. & Schultz, P. G. Adding new chemistries to the genetic code. Annu. Rev. Biochem. 79, 413–444 (2010).

    CAS  PubMed  Google Scholar 

  48. Ryu, Y. & Schultz, P. G. Efficient incorporation of unnatural amino acids into proteins in Escherichia coli. Nat. Methods 3, 263–265 (2006).

    CAS  PubMed  Google Scholar 

  49. Wang, L., Brock, A., Herberich, B. & Schultz, P. G. Expanding the genetic code of Escherichia coli. Science 292, 498–500 (2001).

    CAS  PubMed  Google Scholar 

  50. Tran, A. X., Dong, C. & Whitfield, C. Structure and functional analysis of LptC, a conserved membrane protein involved in the lipopolysaccharide export pathway in Escherichia coli. J. Biol. Chem. 285, 33529–33539 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  51. Bowyer, A., Baardsnes, J., Ajamian, E., Zhang, L. & Cygler, M. Characterization of interactions between LPS transport proteins of the Lpt system. Biochem. Biophys. Res. Commun. 404, 1093–1098 (2011).

    CAS  PubMed  Google Scholar 

  52. Sperandeo, P. et al. New insights into the Lpt machinery for lipopolysaccharide transport to the cell surface: LptA–LptC interaction and LptA stability as sensors of a properly assembled transenvelope complex. J. Bacteriol. 193, 1042–1053 (2011).

    CAS  PubMed  Google Scholar 

  53. Bos, M. P., Tefsen, B., Geurtsen, J. & Tommassen, J. Identification of an outer membrane protein required for the transport of lipopolysaccharide to the bacterial cell surface. Proc. Natl Acad. Sci. USA 101, 9417–9422 (2004).

    CAS  PubMed  Google Scholar 

  54. Braun, M. & Silhavy, T. J. Imp/OstA is required for cell envelope biogenesis in Escherichia coli. Mol. Microbiol. 45, 1289–1302 (2002).

    CAS  PubMed  Google Scholar 

  55. Wu, T. et al. Identification of a protein complex that assembles lipopolysaccharide in the outer membrane of Escherichia coli. Proc. Natl Acad. Sci. USA 103, 11754–11759 (2006).

    CAS  PubMed  Google Scholar 

  56. Hagan, C. L., Silhavy, T. J. & Kahne, D. β-barrel membrane protein assembly by the Bam complex. Annu. Rev. Biochem. 80, 189–210 (2011).

    CAS  PubMed  Google Scholar 

  57. Ruiz, N., Falcone, B., Kahne, D. & Silhavy, T. J. Chemical conditionality: a genetic strategy to probe organelle assembly. Cell 121, 307–317 (2005).

    CAS  PubMed  Google Scholar 

  58. Voulhoux, R., Bos, M. P., Geurtsen, J., Mols, M. & Tommassen, J. Role of a highly conserved bacterial protein in outer membrane protein assembly. Science 299, 262–265 (2003).

    CAS  PubMed  Google Scholar 

  59. Wu, T. et al. Identification of a multicomponent complex required for outer membrane biogenesis in Escherichia coli. Cell 121, 235–245 (2005).

    CAS  PubMed  Google Scholar 

  60. Sklar, J. G. et al. Lipoprotein SmpA is a component of the YaeT complex that assembles outer membrane proteins in Escherichia coli. Proc. Natl Acad. Sci. USA 104, 6400–6405 (2007).

    CAS  PubMed  Google Scholar 

  61. Ricci, D. P. & Silhavy, T. J. The Bam machine: a molecular cooper. Biochim. Biophys. Acta 1818, 1067–1084 (2012).

    CAS  PubMed  Google Scholar 

  62. Rigel, N. W. & Silhavy, T. J. Making a β-barrel: assembly of outer membrane proteins in Gram-negative bacteria. Curr. Opin. Microbiol. 15, 189–193 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  63. Eggert, U. S. et al. Genetic basis for activity differences between vancomycin and glycolipid derivatives of vancomycin. Science 294, 361–364 (2001).

    CAS  PubMed  Google Scholar 

  64. Matsuyama, S., Yokota, N. & Tokuda, H. A novel outer membrane lipoprotein, LolB (HemM), involved in the LolA (p20)-dependent localization of lipoproteins to the outer membrane of Escherichia coli. EMBO J. 16, 6947–6955 (1997).

    CAS  PubMed  PubMed Central  Google Scholar 

  65. Okuda, S. & Tokuda, H. Model of mouth-to-mouth transfer of bacterial lipoproteins through inner membrane LolC, periplasmic LolA, and outer membrane LolB. Proc. Natl Acad. Sci. USA 106, 5877–5882 (2009).

    CAS  PubMed  Google Scholar 

  66. Yakushi, T., Masuda, K., Narita, S., Matsuyama, S. & Tokuda, H. A new ABC transporter mediating the detachment of lipid-modified proteins from membranes. Nat. Cell Biol. 2, 212–218 (2000).

    CAS  PubMed  Google Scholar 

  67. Tsukahara, J., Mukaiyama, K., Okuda, S., Narita, S. & Tokuda, H. Dissection of LolB function — lipoprotein binding, membrane targeting and incorporation of lipoproteins into lipid bilayers. FEBS J. 276, 4496–4504 (2009).

    CAS  PubMed  Google Scholar 

  68. Nakada, S. et al. Structural investigation of the interaction between LolA and LolB using NMR. J. Biol. Chem. 284, 24634–24643 (2009).

    CAS  PubMed  PubMed Central  Google Scholar 

  69. Chng, S. S., Ruiz, N., Chimalakonda, G., Silhavy, T. J. & Kahne, D. Characterization of the two-protein complex in Escherichia coli responsible for lipopolysaccharide assembly at the outer membrane. Proc. Natl Acad. Sci. USA 107, 5363–5368 (2010).

    CAS  PubMed  Google Scholar 

  70. Dong, H. et al. Structural basis for outer membrane lipopolysaccharide insertion. Nature 511, 52–56 (2014).

    CAS  PubMed  Google Scholar 

  71. Qiao, S., Luo, Q., Zhao, Y., Zhang, X. C. & Huang, Y. Structural basis for lipopolysaccharide insertion in the bacterial outer membrane. Nature 511, 108–111 (2014). Together with reference 70, this article reports the first crystal structures of LptDE.

    CAS  PubMed  Google Scholar 

  72. Freinkman, E., Chng, S. S. & Kahne, D. The complex that inserts lipopolysaccharide into the bacterial outer membrane forms a two-protein plug-and-barrel. Proc. Natl Acad. Sci. USA 108, 2486–2491 (2011). This paper establishes that LptD and LptE form a plug-and-barrel structure and suggests that membrane insertion proceeds through a lateral gate in a proline-rich region of the β-barrel of LptD.

    CAS  PubMed  Google Scholar 

  73. Gu, Y. et al. Lipopolysaccharide is inserted into the outer membrane through an intramembrane hole, a lumen gate, and the lateral opening of LptD. Structure 23, 496–504 (2015).

    CAS  PubMed  PubMed Central  Google Scholar 

  74. Li, X., Gu, Y., Dong, H., Wang, W. & Dong, C. Trapped lipopolysaccharide and LptD intermediates reveal lipopolysaccharide translocation steps across the Escherichia coli outer membrane. Sci. Rep. 5, 11883 (2015).

    PubMed  PubMed Central  Google Scholar 

  75. Ahn, V. E. et al. A hydrocarbon ruler measures palmitate in the enzymatic acylation of endotoxin. EMBO J. 23, 2931–2941 (2004). This study is the first to suggest a gating mechanism by which lipids pass into the lumen of a β-barrel through a lateral opening.

    CAS  PubMed  PubMed Central  Google Scholar 

  76. Bishop, R. E. The lipid A palmitoyltransferase PagP: molecular mechanisms and role in bacterial pathogenesis. Mol. Microbiol. 57, 900–912 (2005).

    CAS  PubMed  Google Scholar 

  77. Khan, M. A. & Bishop, R. E. Molecular mechanism for lateral lipid diffusion between the outer membrane external leaflet and a β-barrel hydrocarbon ruler. Biochemistry 48, 9745–9756 (2009).

    CAS  PubMed  PubMed Central  Google Scholar 

  78. Hearn, E. M., Patel, D. R., Lepore, B. W., Indic, M. & van den Berg, B. Transmembrane passage of hydrophobic compounds through a protein channel wall. Nature 458, 367–370 (2009).

    CAS  PubMed  PubMed Central  Google Scholar 

  79. van den Berg, B. Going forward laterally: transmembrane passage of hydrophobic molecules through protein channel walls. Chembiochem 11, 1339–1343 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  80. van den Berg, B., Black, P. N., Clemons, W. M. & Rapoport, T. A. Crystal structure of the long-chain fatty acid transporter FadL. Science 304, 1506–1509 (2004).

    CAS  PubMed  Google Scholar 

  81. Malojcic, G. et al. LptE binds to and alters the physical state of LPS to catalyze its assembly at the cell surface. Proc. Natl Acad. Sci. USA 111, 9467–9472 (2014).

    CAS  PubMed  Google Scholar 

  82. Chng, S. S. et al. Disulfide rearrangement triggered by translocon assembly controls lipopolysaccharide export. Science 337, 1665–1668 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  83. Ruiz, N., Chng, S. S., Hiniker, A., Kahne, D. & Silhavy, T. J. Nonconsecutive disulfide bond formation in an essential integral outer membrane protein. Proc. Natl Acad. Sci. USA 107, 12245–12250 (2010).

    CAS  PubMed  Google Scholar 

  84. Vertommen, D., Ruiz, N., Leverrier, P., Silhavy, T. J. & Collet, J. F. Characterization of the role of the Escherichia coli periplasmic chaperone SurA using differential proteomics. Proteomics 9, 2432–2443 (2009).

    CAS  PubMed  PubMed Central  Google Scholar 

  85. Chimalakonda, G. et al. Lipoprotein LptE is required for the assembly of LptD by the β-barrel assembly machine in the outer membrane of Escherichia coli. Proc. Natl Acad. Sci. USA 108, 2492–2497 (2011).

    CAS  PubMed  Google Scholar 

  86. Denoncin, K., Vertommen, D., Paek, E. & Collet, J. F. The protein–disulfide isomerase DsbC cooperates with SurA and DsbA in the assembly of the essential β-barrel protein LptD. J. Biol. Chem. 285, 29425–29433 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  87. Narita, S., Masui, C., Suzuki, T., Dohmae, N. & Akiyama, Y. Protease homolog BepA (YfgC) promotes assembly and degradation of β-barrel membrane proteins in Escherichia coli. Proc. Natl Acad. Sci. USA 110, E3612–E3621 (2013).

    CAS  PubMed  Google Scholar 

  88. Tran, A. X., Trent, M. S. & Whitfield, C. The LptA protein of Escherichia coli is a periplasmic lipid A-binding protein involved in the lipopolysaccharide export pathway. J. Biol. Chem. 283, 20342–20349 (2008).

    CAS  PubMed  PubMed Central  Google Scholar 

  89. Kaback, H. R. Bacterial membranes. Methods Enzymol. 22, 99–120 (1971).

    Google Scholar 

  90. Kaback, H. R. & Stadtman, E. R. Proline uptake by an isolated cytoplasmic membrane preparation of Escherichia coli. Proc. Natl Acad. Sci. USA 55, 920–927 (1966).

    CAS  PubMed  Google Scholar 

  91. Kim, Y. J., Rajapandi, T. & Oliver, D. SecA protein is exposed to the periplasmic surface of the E. coli inner membrane in its active state. Cell 78, 845–853 (1994).

    CAS  PubMed  Google Scholar 

  92. Masuda, K., Matsuyama, S. & Tokuda, H. Elucidation of the function of lipoprotein-sorting signals that determine membrane localization. Proc. Natl Acad. Sci. USA 99, 7390–7395 (2002).

    CAS  PubMed  Google Scholar 

  93. Ito, Y., Kanamaru, K., Taniguchi, N., Miyamoto, S. & Tokuda, H. A novel ligand bound ABC transporter, LolCDE, provides insights into the molecular mechanisms underlying membrane detachment of bacterial lipoproteins. Mol. Microbiol. 62, 1064–1075 (2006).

    CAS  PubMed  Google Scholar 

  94. Taniguchi, N. & Tokuda, H. Molecular events involved in a single cycle of ligand transfer from an ATP binding cassette transporter, LolCDE, to a molecular chaperone, LolA. J. Biol. Chem. 283, 8538–8544 (2008).

    CAS  PubMed  PubMed Central  Google Scholar 

  95. Mizutani, M. et al. Functional differentiation of structurally similar membrane subunits of the ABC transporter LolCDE complex. FEBS Lett. 587, 23–29 (2013).

    CAS  PubMed  Google Scholar 

  96. Yasuda, M., Iguchi-Yokoyama, A., Matsuyama, S., Tokuda, H. & Narita, S. Membrane topology and functional importance of the periplasmic region of ABC transporter LolCDE. Biosci. Biotechnol. Biochem. 73, 2310–2316 (2009).

    CAS  PubMed  Google Scholar 

  97. Bos, M. P. & Tommassen, J. The LptD chaperone LptE is not directly involved in lipopolysaccharide transport in Neisseria meningitidis. J. Biol. Chem. 286, 28688–28696 (2011).

    CAS  PubMed  PubMed Central  Google Scholar 

  98. Srinivas, N. et al. Peptidomimetic antibiotics target outer-membrane biogenesis in Pseudomonas aeruginosa. Science 327, 1010–1013 (2010).

    CAS  PubMed  Google Scholar 

  99. Werneburg, M. et al. Inhibition of lipopolysaccharide transport to the outer membrane in Pseudomonas aeruginosa by peptidomimetic antibiotics. Chembiochem 13, 1767–1775 (2012).

    CAS  PubMed  Google Scholar 

Download references

Acknowledgements

This work was supported by the National Institute of Allergy and Infectious Diseases (AI081059 to D.K.) and the National Institute of General Medical Sciences (GM034821 to T.J.S.; GM100951 to N.R.), under the US National Institutes of Health (NIH).

Author information

Authors and Affiliations

  1. Department of Chemistry and Chemical Biology, Harvard University, Cambridge, 02138, Massachusetts, USA

    Suguru Okuda, David J. Sherman & Daniel Kahne

  2. Department of Bio-system Pharmacology, Graduate School of Medicine, Osaka University, Osaka, 565–0871, Japan

    Suguru Okuda

  3. Department of Molecular Biology, Princeton University, Princeton, 08544, New Jersey, USA

    Thomas J. Silhavy

  4. Department of Microbiology, The Ohio State University, Columbus, 43210, Ohio, USA

    Natividad Ruiz

  5. Department of Molecular and Cellular Biology, Harvard University, Cambridge, 02138, Massachusetts, USA

    Daniel Kahne

  6. Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, 02115, Massachusetts, USA

    Daniel Kahne

Authors
  1. Suguru Okuda
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. David J. Sherman
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Thomas J. Silhavy
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Natividad Ruiz
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Daniel Kahne
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Daniel Kahne.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Related links

PowerPoint slides

Glossary

Periplasm

An aqueous, densely packed compartment between the Gram-negative inner membrane and outer membrane. The periplasm has a unique assortment of proteins and also contains a thin layer of peptidoglycan.

Lipid A

The hydrophobic glucosamine-based phospholipid anchor of lipopolysaccharide molecules. Lipid A is also known as endotoxin.

O antigen

Attached to the core oligosaccharide, this repetitive glycan is the outermost part of the lipopolysaccharide molecule and is a target of the host immune system.

ATP-binding cassette transporter

(ABC transporter). A transmembrane protein complex that uses the energy derived from ATP binding and hydrolysis to transport various substrates. These proteins are members of one of the largest protein superfamilies and consist of transmembrane domains and conserved nucleotide-binding domains.

Bitopic membrane protein

A type of membrane protein that contains only one transmembrane helix.

β-barrel

A class of integral membrane protein comprised of β-strands that satisfy their peptide backbone hydrogen bonds by forming a cylindrical barrel structure, which exposes hydrophobic side chains to the membrane and shields hydrophilic side chains.

Lipoprotein

A protein characterized by the presence of an amino-terminal lipid-modified cysteine that anchors the hydrophilic protein to the cell membrane.

Transposon mutants

Mutants that are created through the random insertion of a transposon (or transposable element) into a genome. In the cited study, the transposable element encoded an arabinose-inducible promoter that could drive the expression of chromosomal genes located immediately downstream of the transposon insertion site. It was used to identify essential genes by screening for transposon mutants that required the presence of arabinose in the medium for growth.

Synteny

The colocalization of genes in a genetic locus.

Reductionist bioinformatics approach

An approach to bioinformatics in which a complex biological system is studied through a comparative analysis of similar simpler systems. In this case, a comparative bioinformatics search was used to decrease the number of candidate genes of interest by comparing the genome of interest with other genomes of smaller size.

β-jellyroll

A structure in which anti-parallel β-strands are 'wrapped' into a cylindrical, barrel-like shape without necessarily maintaining a continuous hydrogen bonding network.

Periplasmic chaperone

A periplasmic protein that prevents macromolecules from aggregating and assists them in reaching their destination.

Spheroplasts

Osmotically fragile bacterial cells that have had their outer membranes and peptidoglycan layers incompletely disrupted, which causes them to form a spherical shape.

Photo-crosslinking

The light-induced formation of a covalent bond between two molecules to detect molecular interactions.

Unnatural amino acid

Non-coded, non-proteinogenic amino acid that, when incorporated into proteins, enables various new functions.

Size-exclusion chromatography

A chromatographic technique that is used for preparative or analytical purposes to separate molecules (usually macromolecules) based on their size.

β-barrel assembly machine pathway

(Bam pathway). Following secretion from the inner membrane and translocation across the periplasm, the Bam complex is responsible for folding and inserting β-barrel proteins into the outer membrane. In Escherichia coli, the Bam complex is composed of one β-barrel protein, BamA, and four outer membrane lipoproteins, BamB, BamC, BamD and BamE.

Localization of lipoproteins pathway

(Lol pathway). A chaperone-based transport pathway that is involved in transporting outer membrane lipoproteins from the outer leaflet of the inner membrane to the inner leaflet of the outer membrane.

Trypsin

A serine protease that hydrolyses peptide bonds on the carboxy-terminal side of lysine and arginine residues, and is commonly used to determine the stability of proteins.

Crenellated β-barrel

A β-barrel protein in which the formation of inter-strand hydrogen bonds is disrupted, creating openings similar to the crenels in the turret of a castle.

Right-side-out membrane vesicles

Membrane vesicles with a native orientation that are prepared by the osmotic lysis of spheroplasts.

Vanadate

Sodium orthovanadate (Na3VO4) inhibits protein tyrosine phosphatases, alkaline phosphatases and many ATPases by acting as a phosphate analogue and binding in the active site in which phosphate usually binds.

Peptidomimetic

Chemical compounds that mimic natural peptides because of their small protein-like chains.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Okuda, S., Sherman, D., Silhavy, T. et al. Lipopolysaccharide transport and assembly at the outer membrane: the PEZ model. Nat Rev Microbiol 14, 337–345 (2016). https://doi.org/10.1038/nrmicro.2016.25

Download citation

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nrmicro.2016.25

This article is cited by

Search

Advanced search

Quick links

Nature Briefing Microbiology

Sign up for the Nature Briefing: Microbiology newsletter — what matters in microbiology research, free to your inbox weekly.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing: Microbiology