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.

  • Article
  • Published:

Insights into noncanonical E1 enzyme activation from the structure of autophagic E1 Atg7 with Atg8

Nature Structural & Molecular Biology volume 18, pages 1323–1330 (2011)Cite this article

Subjects

Abstract

Autophagy is the degradation of cellular organelles via the lysosomal pathway. The autophagic ubiquitin-like (Ubl) molecule Atg8 is activated by the E1-like enzyme Atg7. As this noncanonical E1 enzyme's domain organization is unique among Ubl-activating E1 enzymes, the structural basis for its interactions with Atg8 and partner E2 enzymes remains obscure. Here we present the structure of the N-terminal domain of Atg7, revealing a unique protein fold and interactions with both autophagic E2 enzymes Atg3 and Atg10. The structure of the C-terminal domain of Atg7 in complex with Atg8 shows the mode of dimerization and mechanism of recognition of Atg8. Notably, the catalytic cysteine residue in Atg7 is positioned close to the C-terminal glycine of Atg8, its target for thioester formation, potentially eliminating the need for large conformational rearrangements characteristic of other E1s.

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: Structure of Atg7C−Atg8 complex and formation of Atg7C~Atg8 conjugate.
Figure 2: Atg8 recognition by the C-terminal region of Atg7C.
Figure 3: Active site of Atg7C−Atg8 complex.
Figure 4: Interaction between yeast Atg7 and human LC3B mutant.
Figure 5: Structure of Atg7N and its interaction with autophagic E2 enzymes.
Figure 6: Interaction map of Atg7 with other autophagic molecules and a model for the Atg3−Atg7−Atg8 ternary complex.

Similar content being viewed by others

Accession codes

Primary accessions

Protein Data Bank

Referenced accessions

Protein Data Bank

References

  1. Nakatogawa, H., Suzuki, K., Kamada, Y. & Ohsumi, Y. Dynamics and diversity in autophagy mechanisms: lessons from yeast. Nat. Rev. Mol. Cell Biol. 10, 458–467 (2009).

    Article  CAS  PubMed  Google Scholar 

  2. Klionsky, D.J. & Emr, S.D. Autophagy as a regulated pathway of cellular degradation. Science 290, 1717–1721 (2000).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Klionsky, D.J. et al. A unified nomenclature for yeast autophagy-related genes. Dev. Cell 5, 539–545 (2003).

    Article  CAS  PubMed  Google Scholar 

  4. Noda, N.N., Ohsumi, Y. & Inagaki, F. ATG systems from the protein structural point of view. Chem. Rev. 109, 1587–1598 (2009).

    Article  CAS  PubMed  Google Scholar 

  5. Suzuki, K. & Ohsumi, Y. Current knowledge of the pre-autophagosomal structure (PAS). FEBS Lett. 584, 1280–1286 (2010).

    Article  CAS  PubMed  Google Scholar 

  6. Shintani, T. & Klionsky, D.J. Autophagy in health and disease: a double-edged sword. Science 306, 990–995 (2004).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Rabinowitz, J.D. & White, E. Autophagy and metabolism. Science 330, 1344–1348 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Klionsky, D.J. et al. A comprehensive glossary of autophagy-related molecules and processes. Autophagy 6, 438–448 (2010).

    Article  PubMed  Google Scholar 

  9. Levine, B., Mizushima, N. & Virgin, H.W. Autophagy in immunity and inflammation. Nature 469, 323–335 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Mizushima, N. et al. A protein conjugation system essential for autophagy. Nature 395, 395–398 (1998).

    Article  CAS  PubMed  Google Scholar 

  11. Ichimura, Y. et al. A ubiquitin-like system mediates protein lipidation. Nature 408, 488–492 (2000).

    Article  CAS  PubMed  Google Scholar 

  12. Varshavsky, A. Regulated protein degradation. Trends Biochem. Sci. 30, 283–286 (2005).

    Article  CAS  PubMed  Google Scholar 

  13. Hochstrasser, M. Origin and function of ubiquitin-like proteins. Nature 458, 422–429 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Hershko, A. & Ciechanover, A. The ubiquitin system. Annu. Rev. Biochem. 67, 425–479 (1998).

    Article  CAS  PubMed  Google Scholar 

  15. Schulman, B.A. & Harper, J.W. Ubiquitin-like protein activation by E1 enzymes: the apex for downstream signalling pathways. Nat. Rev. Mol. Cell Biol. 10, 319–331 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Lee, I. & Schindelin, H. Structural insights into E1-catalyzed ubiquitin activation and transfer to conjugating enzymes. Cell 134, 268–278 (2008).

    Article  CAS  PubMed  Google Scholar 

  17. Huang, D.T. et al. Basis for a ubiquitin-like protein thioester switch toggling E1–E2 affinity. Nature 445, 394–398 (2007).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Olsen, S.K., Capili, A.D., Lu, X., Tan, D.S. & Lima, C.D. Active site remodelling accompanies thioester bond formation in the SUMO E1. Nature 463, 906–912 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Völler, D. & Schindelin, H. And yet it moves: active site remodeling in the SUMO E1. Structure 18, 419–421 (2010).

    Article  PubMed  Google Scholar 

  20. Bacik, J.P., Walker, J.R., Ali, M., Schimmer, A.D. & Dhe-Paganon, S. Crystal structure of the human ubiquitin-activating enzyme 5 (UBA5) bound to ATP: mechanistic insights into a minimalistic E1 enzyme. J. Biol. Chem. 285, 20273–20280 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Geng, J. & Klionsky, D.J. The Atg8 and Atg12 ubiquitin-like conjugation systems in macroautophagy. EMBO Rep. 9, 859–864 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Zhang, J. & Ney, P.A. Reticulocyte mitophagy: monitoring mitochondrial clearance in a mammalian model. Autophagy 6, 405–408 (2010).

    Article  CAS  PubMed  Google Scholar 

  23. Komatsu, M. et al. Impairment of starvation-induced and constitutive autophagy in Atg7-deficient mice. J. Cell Biol. 169, 425–434 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. Mortensen, M. et al. The autophagy protein Atg7 is essential for hematopoietic stem cell maintenance. J. Exp. Med. 208, 455–467 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Lenz, H.D., Vierstra, R.D., Nurnberger, T. & Gust, A.A. ATG7 contributes to plant basal immunity towards fungal infection. Plant Signal. Behav. 6, 1040–1042 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Komatsu, M. et al. The C-terminal region of an Apg7p/Cvt2p is required for homodimerization and is essential for its E1 activity and E1–E2 complex formation. J. Biol. Chem. 276, 9846–9854 (2001).

    Article  CAS  PubMed  Google Scholar 

  27. Noda, T., Fujita, N. & Yoshimori, T. The Ubi brothers reunited. Autophagy 4, 540–541 (2008).

    Article  CAS  PubMed  Google Scholar 

  28. Holm, L. & Rosenstrom, P. Dali server: conservation mapping in 3D. Nucleic Acids Res. 38, W545–W549 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Yamazaki-Sato, H., Tanida, I., Ueno, T. & Kominami, E. The carboxyl terminal 17 amino acids within Apg7 are essential for Apg8 lipidation, but not for Apg12 conjugation. FEBS Lett. 551, 71–77 (2003).

    Article  CAS  PubMed  Google Scholar 

  30. Lake, M.W., Wuebbens, M.M., Rajagopalan, K.V. & Schindelin, H. Mechanism of ubiquitin activation revealed by the structure of a bacterial MoeB-MoaD complex. Nature 414, 325–329 (2001).

    Article  CAS  PubMed  Google Scholar 

  31. Lehmann, C., Begley, T.P. & Ealick, S.E. Structure of the Escherichia coli ThiS-ThiF complex, a key component of the sulfur transfer system in thiamin biosynthesis. Biochemistry 45, 11–19 (2006).

    Article  CAS  PubMed  Google Scholar 

  32. Lois, L.M. & Lima, C.D. Structures of the SUMO E1 provide mechanistic insights into SUMO activation and E2 recruitment to E1. EMBO J. 24, 439–451 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Harding, M.M. Small revisions to predicted distances around metal sites in proteins. Acta Crystallogr. D Biol. Crystallogr. 62, 678–682 (2006).

    Article  PubMed  Google Scholar 

  34. Reynolds, C., Damerell, D. & Jones, S. ProtorP: a protein-protein interaction analysis server. Bioinformatics 25, 413–414 (2009).

    Article  CAS  PubMed  Google Scholar 

  35. Hurley, J.H., Lee, S. & Prag, G. Ubiquitin-binding domains. Biochem. J. 399, 361–372 (2006).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. Sato, Y. et al. Structural basis for specific cleavage of Lys 63-linked polyubiquitin chains. Nature 455, 358–362 (2008).

    Article  CAS  PubMed  Google Scholar 

  37. Yamada, Y. et al. The crystal structure of Atg3, an autophagy-related ubiquitin carrier protein (E2) enzyme that mediates Atg8 lipidation. J. Biol. Chem. 282, 8036–8043 (2007).

    Article  CAS  PubMed  Google Scholar 

  38. Yamaguchi, M. et al. Autophagy-related protein 8 (Atg8) family interacting motif in Atg3 mediates the Atg3-Atg8 interaction and is crucial for the cytoplasm-to-vacuole targeting pathway. J. Biol. Chem. 285, 29599–29607 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  39. Noda, N.N., Ohsumi, Y. & Inagaki, F. Atg8-family interacting motif crucial for selective autophagy. FEBS Lett. 584, 1379–1385 (2010).

    Article  CAS  PubMed  Google Scholar 

  40. Ichimura, Y. et al. Structural basis for sorting mechanism of p62 in selective autophagy. J. Biol. Chem. 283, 22847–22857 (2008).

    Article  CAS  PubMed  Google Scholar 

  41. Nakatogawa, H., Ichimura, Y. & Ohsumi, Y. Atg8, a ubiquitin-like protein required for autophagosome formation, mediates membrane tethering and hemifusion. Cell 130, 165–178 (2007).

    Article  CAS  PubMed  Google Scholar 

  42. Suzuki, K., Kubota, Y., Sekito, T. & Ohsumi, Y. Hierarchy of Atg proteins in pre-autophagosomal structure organization. Genes Cells 12, 209–218 (2007).

    Article  CAS  PubMed  Google Scholar 

  43. Huang, W.P., Scott, S.V., Kim, J. & Klionsky, D.J. The itinerary of a vesicle component, Aut7p/Cvt5p, terminates in the yeast vacuole via the autophagy/Cvt pathways. J. Biol. Chem. 275, 5845–5851 (2000).

    Article  CAS  PubMed  Google Scholar 

  44. Hanada, T. et al. The Atg12-Atg5 conjugate has a novel E3-like activity for protein lipidation in autophagy. J. Biol. Chem. 282, 37298–37302 (2007).

    Article  CAS  PubMed  Google Scholar 

  45. Otwinowski, Z. & Minor, W. Processing of X-ray diffraction data collected in oscillation mode. Methods Enzymol. 276, 307–326 (1997).

    Article  CAS  PubMed  Google Scholar 

  46. Adams, P.D. et al. PHENIX: a comprehensive Python-based system for macromolecular structure solution. Acta Crystallogr. D Biol. Crystallogr. 66, 213–221 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  47. Emsley, P. & Cowtan, K. Coot: model-building tools for molecular graphics. Acta Crystallogr. D Biol. Crystallogr. 60, 2126–2132 (2004).

    Article  PubMed  Google Scholar 

  48. Jones, T.A., Zou, J.-Y., Cowan, S.W. & Kjeldgaard, M. Improved methods for binding protein models in electron density maps and the location of errors in these models. Acta Crystallogr. A 47, 110–119 (1991).

    Article  PubMed  Google Scholar 

  49. Chen, V.B. et al. MolProbity: all-atom structure validation for macromolecular crystallography. Acta Crystallogr. D Biol. Crystallogr. 66, 12–21 (2010).

    Article  CAS  PubMed  Google Scholar 

  50. Heinig, M. & Frishman, D. STRIDE: a web server for secondary structure assignment from known atomic coordinates of proteins. Nucleic Acids Res. 32, W500–W502 (2004).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

We thank the staff at 4A beamline, Pohang Accelerator Laboratory, South Korea, and NE3-A and NW12 beamline, Photon Factory, Japan, for help with the data collection; H. Nakatogawa and Y. Ohsumi (Tokyo Institute of Technology), W.-K. Huh (Seoul National University), C.-W. Yun (Korea University), D.J. Klionsky (University of Michigan) for yeast strains; and M.J. Eck for critical comments on the manuscript. This work was supported by the World-Class University Project (R33-10108), the 21C Frontier Functional Proteomics Project (FPR08B2-270) and the Korea Healthcare Technology R&D Project, Ministry for Health, Welfare & Family Affairs, Republic of Korea (A092006 and A084016).

Author information

Authors and Affiliations

  1. School of Life Sciences and Biotechnology, Korea University, Seoul, South Korea

    Seung Beom Hong, Byeong-Won Kim, Se Woong Kim, Joon Kim & Hyun Kyu Song

  2. Biomedical Research Center, Korea Institute of Science and Technology, Seoul, South Korea

    Kyung-Eun Lee & Hyesung Jeon

Authors
  1. Seung Beom Hong
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Byeong-Won Kim
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Kyung-Eun Lee
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Se Woong Kim
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Hyesung Jeon
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Joon Kim
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Hyun Kyu Song
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

S.B.H., B.-W.K. and H.K.S. performed biochemical and structural studies; S.W.K. and J.K. performed yeast genetics; K.-E.L. and H.J. performed EM studies; S.B.H. and H.K.S. designed experiments, analyzed data and wrote the manuscript.

Corresponding author

Correspondence to Hyun Kyu Song.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Text and Figures

Supplementary Figs. 1–8, Supplementary Tables 1 and 2 and Supplementary Methods (PDF 2589 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Hong, S., Kim, BW., Lee, KE. et al. Insights into noncanonical E1 enzyme activation from the structure of autophagic E1 Atg7 with Atg8. Nat Struct Mol Biol 18, 1323–1330 (2011). https://doi.org/10.1038/nsmb.2165

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nsmb.2165

Search

Advanced search

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