Skip to main content

Thank you for visiting 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.

Structural basis of enzyme encapsulation into a bacterial nanocompartment


Compartmentalization is an important organizational feature of life. It occurs at varying levels of complexity ranging from eukaryotic organelles and the bacterial microcompartments, to the molecular reaction chambers formed by enzyme assemblies. The structural basis of enzyme encapsulation in molecular compartments is poorly understood. Here we show, using X-ray crystallographic, biochemical and EM experiments, that a widespread family of conserved bacterial proteins, the linocin-like proteins, form large assemblies that function as a minimal compartment to package enzymes. We refer to this shell-forming protein as 'encapsulin'. The crystal structure of such a particle from Thermotoga maritima determined at 3.1-Å resolution reveals that 60 copies of the monomer assemble into a thin, icosahedral shell with a diameter of 240 Å. The interior of this nanocompartment is lined with conserved binding sites for short polypeptide tags present as C-terminal extensions of enzymes involved in oxidative-stress response.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.


All prices are NET prices.

Figure 1: Alignment of selected encapsulin proteins.
Figure 2: Structure of the T. maritima encapsulin.
Figure 3: Structural comparison of T. maritima encapsulin with P. furiosus PfV and gp5 of the HK97 virus.
Figure 4: Packaged proteins possess a C-terminal extension.
Figure 5: Packaging of oligomeric cargo proteins.
Figure 6: Schematic architectural and primary-structure organization of the different types of encapsulins.

Accession codes

Primary accessions

Protein Data Bank

Referenced accessions

Protein Data Bank


  1. 1

    Cannon, G.C. et al. Microcompartments in prokaryotes: carboxysomes and related polyhedra. Appl. Environ. Microbiol. 67, 5351–5361 (2001).

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  2. 2

    Kerfeld, C.A. et al. Protein structures forming the shell of primitive bacterial organelles. Science 309, 936–938 (2005).

    CAS  Article  PubMed  Google Scholar 

  3. 3

    Tanaka, S. et al. Atomic-level models of the bacterial carboxysome shell. Science 319, 1083–1086 (2008).

    CAS  Article  PubMed  Google Scholar 

  4. 4

    Havemann, G.D., Sampson, E.M. & Bobik, T.A. PduA is a shell protein of polyhedral organelles involved in coenzyme B12-dependent degradation of 1,2-propanediol in Salmonella enterica serovar typhimurium LT2. J. Bacteriol. 184, 1253–1261 (2002).

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  5. 5

    Kofoid, E., Rappleye, C., Stojiljkovic, I. & Roth, J. The 17-gene ethanolamine (eut) operon of Salmonella typhimurium encodes five homologues of carboxysome shell proteins. J. Bacteriol. 181, 5317–5329 (1999).

    CAS  PubMed  PubMed Central  Google Scholar 

  6. 6

    Anderson, D.H., Kickhoefer, V.A., Sievers, S.A., Rome, L.H. & Eisenberg, D. Draft crystal structure of the vault shell at 9- resolution. PLoS Biol. 5, e318 (2007).

    Article  PubMed  PubMed Central  Google Scholar 

  7. 7

    Smith, J.L. The physiological role of ferritin-like compounds in bacteria. Crit. Rev. Microbiol. 30, 173–185 (2004).

    CAS  Article  PubMed  Google Scholar 

  8. 8

    Ævarsson, A., Seger, K., Turley, S., Sokatch, J.R. & Hol, W.G. Crystal structure of 2-oxoisovalerate and dehydrogenase and the architecture of 2-oxo acid dehydrogenase multienzyme complexes. Nat. Struct. Biol. 6, 785–792 (1999).

    Article  PubMed  Google Scholar 

  9. 9

    Ritsert, K. et al. Studies on the lumazine synthase/riboflavin synthase complex of Bacillus subtilis: crystal structure analysis of reconstituted, icosahedral β-subunit capsids with bound substrate analogue inhibitor at 2.4 resolution. J. Mol. Biol. 253, 151–167 (1995).

    CAS  Article  PubMed  Google Scholar 

  10. 10

    Jenni, S. et al. Structure of fungal fatty acid synthase and implications for iterative substrate shuttling. Science 316, 254–261 (2007).

    CAS  Article  PubMed  Google Scholar 

  11. 11

    Valdes-Stauber, N. & Scherer, S. Isolation and characterization of Linocin M18, a bacteriocin produced by Brevibacterium linens. Appl. Environ. Microbiol. 60, 3809–3814 (1994).

    CAS  PubMed  PubMed Central  Google Scholar 

  12. 12

    Hicks, P.M., Rinker, K.D., Baker, J.R. & Kelly, R.M. Homomultimeric protease in the hyperthermophilic bacterium Thermotoga maritima has structural and amino acid sequence homology to bacteriocins in mesophilic bacteria. FEBS Lett. 440, 393–398 (1998).

    CAS  Article  PubMed  Google Scholar 

  13. 13

    Rosenkrands, I. et al. Identification and characterization of a 29-kilodalton protein from Mycobacterium tuberculosis culture filtrate recognized by mouse memory effector cells. Infect. Immun. 66, 2728–2735 (1998).

    CAS  PubMed  PubMed Central  Google Scholar 

  14. 14

    Wikoff, W.R. et al. Topologically linked protein rings in the bacteriophage HK97 capsid. Science 289, 2129–2133 (2000).

    CAS  Article  PubMed  Google Scholar 

  15. 15

    Akita, F. et al. The crystal structure of a virus-like particle from the hyperthermophilic archaeon Pyrococcus furiosus provides insight into the evolution of viruses. J. Mol. Biol. 368, 1469–1483 (2007).

    CAS  Article  PubMed  Google Scholar 

  16. 16

    Helgstrand, C. et al. The refined structure of a protein catenane: the HK97 bacteriophage capsid at 3.44 resolution. J. Mol. Biol. 334, 885–899 (2003).

    CAS  Article  PubMed  Google Scholar 

  17. 17

    Altschul, S.F., Gish, W., Miller, W., Myers, E.W. & Lipman, D.J. Basic local alignment search tool. J. Mol. Biol. 215, 403–410 (1990).

    CAS  Article  PubMed  Google Scholar 

  18. 18

    Altschul, S.F. et al. Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res. 25, 3389–3402 (1997).

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  19. 19

    Kim, S.J. & Shoda, M. Purification and characterization of a novel peroxidase from Geotrichum candidum Dec 1 involved in decolorization of dyes. Appl. Environ. Microbiol. 65, 1029–1035 (1999).

    CAS  PubMed  PubMed Central  Google Scholar 

  20. 20

    Chang, C., Evdokimova, E., Savchenko, A. & Joachimiak, A. & Midwest Center for Structural Genomics. Crystal structure of protein NE0167 from Nitrosomonas europaea, PDB 1ZPY, doi:102210/pdb1zpy/pdb (2005).

  21. 21

    Marcotte, E.M. et al. Detecting protein function and protein-protein interactions from genome sequences. Science 285, 751–753 (1999).

    CAS  Article  PubMed  Google Scholar 

  22. 22

    Valdes-Stauber, N. & Scherer, S. Nucleotide sequence and taxonomical distribution of the bacteriocin gene lin cloned from Brevibacterium linens M18. Appl. Environ. Microbiol. 62, 1283–1286 (1996).

    CAS  PubMed  PubMed Central  Google Scholar 

  23. 23

    Zubieta, C. et al. Crystal structures of two novel dye-decolorizing peroxidases reveal a β-barrel fold with a conserved heme-binding motif. Proteins 69, 223–233 (2007).

    CAS  Article  PubMed  Google Scholar 

  24. 24

    Bamford, D.H., Grimes, J.M. & Stuart, D.I. What does structure tell us about virus evolution? Curr. Opin. Struct. Biol. 15, 655–663 (2005).

    CAS  Article  PubMed  Google Scholar 

  25. 25

    Takahashi, T. & Kuyucak, S. Functional properties of threefold and fourfold channels in ferritin deduced from electrostatic calculations. Biophys. J. 84, 2256–2263 (2003).

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  26. 26

    Harrison, P.M. & Arosio, P. The ferritins: molecular properties, iron storage function and cellular regulation. Biochim. Biophys. Acta 1275, 161–203 (1996).

    Article  PubMed  Google Scholar 

  27. 27

    Rocha, E.R., Andrews, S.C., Keen, J.N. & Brock, J.H. Isolation of a ferritin from Bacteroides fragilis. FEMS Microbiol. Lett. 95, 207–212 (1992).

    CAS  Article  Google Scholar 

  28. 28

    Finegold, S.M. & George, W.L. Anaerobic Infections in Humans Vol. XXI 851 (Academic Press, San Diego, CA, 1989).

  29. 29

    Manca, C., Paul, S., Barry, C.E. III, Freedman, V.H. & Kaplan, G. Mycobacterium tuberculosis catalase and peroxidase activities and resistance to oxidative killing in human monocytes in vitro. Infect. Immun. 67, 74–79 (1999).

    CAS  PubMed  PubMed Central  Google Scholar 

  30. 30

    Tsai, Y. et al. Structural analysis of CsoS1A and the protein shell of the Halothiobacillus neapolitanus carboxysome. PLoS Biol. 5, e144 (2007).

    Article  PubMed  PubMed Central  Google Scholar 

  31. 31

    Yeates, T.O., Tsai, Y., Tanaka, S., Sawaya, M.R. & Kerfeld, C.A. Self-assembly in the carboxysome: a viral capsid-like protein shell in bacterial cells. Biochem. Soc. Trans. 35, 508–511 (2007).

    CAS  Article  PubMed  Google Scholar 

  32. 32

    Tobimatsu, T., Kawata, M. & Toraya, T. The N-terminal regions of β and γ subunits lower the solubility of adenosylcobalamin-dependent diol dehydratase. Biosci. Biotechnol. Biochem. 69, 455–462 (2005).

    CAS  Article  PubMed  Google Scholar 

  33. 33

    Stauber, N.V. Isolierung, Charakterisierung und Nukleinsäuresequenz eines Bakteriozins aus Brevibacterium linens M18. Doktor der Naturwissenschaften Thesis, Technische Universität München-Weihenstephan, (1995).

  34. 34

    Seebeck, F.P., Woycechowsky, K.J., Zhuang, W., Rabe, J.P. & Hilvert, D. A simple tagging system for protein encapsulation. J. Am. Chem. Soc. 128, 4516–4517 (2006).

    CAS  Article  PubMed  Google Scholar 

  35. 35

    Huber, R. et al. Thermotoga maritima sp. nov represents a new genus of unique extremely thermophilic eubacteria growing up to 90 °C. Arch. Microbiol. 144, 324–333 (1986).

    CAS  Article  Google Scholar 

  36. 36

    Santimone, M. Titration study of guaiacol oxidation by horseradish peroxidase. Can. J. Biochem. 53, 649–657 (1975).

    CAS  Article  PubMed  Google Scholar 

  37. 37

    Kabsch, W. Automatic processing of rotation diffraction data from crystals of initially unknown symmetry and cell constants. J. Appl. Crystallogr. 26, 795–800 (1993).

    CAS  Article  Google Scholar 

  38. 38

    Rossmann, M.G. & Blow, D.M. The detection of sub-units within the crystallographic asymmetric unit. Acta Crystallogr. 15, 24–31 (1962).

    CAS  Article  Google Scholar 

  39. 39

    Tong, L. & Rossmann, M.G. Rotation function calculations with GLRF program. Methods Enzymol. 276, 594–611 (1997).

    CAS  Article  PubMed  Google Scholar 

  40. 40

    Kleywegt, G.J. & Read, R.J. Not your average density. Structure 5, 1557–1569 (1997).

    CAS  Article  PubMed  Google Scholar 

  41. 41

    Cowtan, K.D. & Main, P. Improvement of macromolecular electron-density maps by the simultaneous application of real and reciprocal space constraints. Acta Crystallogr. D Biol. Crystallogr. 49, 148–157 (1993).

    CAS  Article  PubMed  Google Scholar 

  42. 42

    Murshudov, G.N., Vagin, A.A. & Dodson, E.J. Refinement of macromolecular structures by the maximum-likelihood method. Acta Crystallogr. D Biol. Crystallogr. 53, 240–255 (1997).

    CAS  Article  PubMed  Google Scholar 

  43. 43

    Afonine, P.V., Grosse-Kunstleve, R.W. & Adams, P.D. CCP4 Newsletter No. 42, Contribution 8, (Daresbury Laboratory, Warrington, Cheshire, UK, 2005).

  44. 44

    Krissinel, E. & Henrick, K. Secondary-structure matching (SSM), a new tool for fast protein structure alignment in three dimensions. Acta Crystallogr. D Biol. Crystallogr. 60, 2256–2268 (2004).

    CAS  Article  PubMed  Google Scholar 

  45. 45

    Ohi, M., Li, Y., Cheng, Y. & Walz, T. Negative staining and image classification—powerful tools in modern electron microscopy. Biol. Proced. Online 6, 23–34 (2004).

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  46. 46

    Nickell, S. et al. TOM software toolbox: acquisition and analysis for electron tomography. J. Struct. Biol. 149, 227–234 (2005).

    Article  PubMed  Google Scholar 

  47. 47

    Sander, B., Golas, M.M. & Stark, H. Automatic CTF correction for single particles based upon multivariate statistical analysis of individual power spectra. J. Struct. Biol. 142, 392–401 (2003).

    CAS  Article  PubMed  Google Scholar 

  48. 48

    van Heel, M., Harauz, G., Orlova, E.V., Schmidt, R. & Schatz, M. A new generation of the IMAGIC image processing system. J. Struct. Biol. 116, 17–24 (1996).

    CAS  Article  PubMed  Google Scholar 

  49. 49

    Dube, P., Tavares, P., Lurz, R. & van Heel, M. The portal protein of bacteriophage SPP1: a DNA pump with 13-fold symmetry. EMBO J. 12, 1303–1309 (1993).

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  50. 50

    van Heel, M. & Frank, J. Use of multivariate statistics in analysing the images of biological macromolecules. Ultramicroscopy 6, 187–194 (1981).

    CAS  PubMed  Google Scholar 

  51. 51

    van Heel, M. Angular reconstitution: a posteriori assignment of projection directions for 3D reconstruction. Ultramicroscopy 21, 111–123 (1987).

    CAS  Article  PubMed  Google Scholar 

  52. 52

    Orlova, E.V. et al. Structure of keyhole limpet hemocyanin type 1 (KLH1) at 15 resolution by electron cryomicroscopy and angular reconstitution. J. Mol. Biol. 271, 417–437 (1997).

    CAS  Article  PubMed  Google Scholar 

  53. 53

    Sali, A. & Blundell, T.L. Comparative protein modelling by satisfaction of spatial restraints. J. Mol. Biol. 234, 779–815 (1993).

    CAS  Article  PubMed  Google Scholar 

  54. 54

    Wriggers, W., Milligan, R.A. & McCammon, J.A. Situs: a package for docking crystal structures into low-resolution maps from electron microscopy. J. Struct. Biol. 125, 185–195 (1999).

    CAS  Article  PubMed  Google Scholar 

  55. 55

    Barton, G.J. ALSCRIPT: a tool to format multiple sequence alignments. Protein Eng. 6, 37–40 (1993).

    CAS  Article  PubMed  Google Scholar 

Download references


Crystallographic data were collected at the beamline X06SA at the Swiss Light Source (SLS). We are grateful to C. Schulze-Briese, E. Pohl and T. Tomizaki for their outstanding support at the SLS. We thank R. Brunisholz, P. Hunziker and Y. Auchli at the Functional Genomics Center Zurich for mass-spectrometric analysis, the Electron Microscopy Center Zurich (EMEZ) and Martin Beck for support with EM data collection, M. Müller, F. Voigts-Hoffmann and F. Imkamp for critically reading the manuscript and all members of the Ban and Weber-Ban laboratory for suggestions and discussions. D.B. was supported by a Federation of European Biochemical Societies long-term fellowship. This work was supported by the Swiss National Science Foundation (SNSF) (to N.B. and E.W.B.) and the National Center of Excellence in Research (NCCR) Structural Biology program of the SNSF (to N.B. and E.W.B.).

Author information



Corresponding authors

Correspondence to Eilika Weber-Ban or Nenad Ban.

Ethics declarations

Competing interests

The authors are planning on filing a pre-patent application on possible uses and applications of encapsulin in biotechnology and biomedicine.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–3 and Supplementary data (PDF 1461 kb)

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Sutter, M., Boehringer, D., Gutmann, S. et al. Structural basis of enzyme encapsulation into a bacterial nanocompartment. Nat Struct Mol Biol 15, 939–947 (2008).

Download citation

Further reading


Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

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