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SUMO modification of the ubiquitin-conjugating enzyme E2-25K

Nature Structural & Molecular Biology volume 12, pages 264–269 (2005)Cite this article

Abstract

Post-translational modification with small ubiquitin-related modifier (SUMO) alters the function of many proteins, but the molecular mechanisms and consequences of this modification are still poorly defined. During a screen for novel SUMO1 targets, we identified the ubiquitin-conjugating enzyme E2-25K (Hip2). SUMO attachment severely impairs E2-25K ubiquitin thioester and unanchored ubiquitin chain formation in vitro. Crystal structures of E2-25K(1–155) and of the E2-25K(1–155)–SUMO conjugate (E2-25K*SUMO) indicate that SUMO attachment interferes with E1 interaction through its location on the N-terminal helix. The SUMO acceptor site in E2-25K, Lys14, does not conform to the consensus site found in most SUMO targets (ΨKXE), and functions only in the context of an α-helix. In contrast, adjacent SUMO consensus sites are modified only when in unstructured peptides. The demonstration that secondary structure elements are part of SUMO attachment signals could contribute to a better prediction of SUMO targets.

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Figure 1: E2-25K is SUMOylated in vitro and in vivo.
Figure 2: Sumoylation of E2-25K inhibits ubiquitin thioester formation.
Figure 3: Crystal structure of SUMO-modified E2-25K.
Figure 4: SUMO target sites are defined by their structural context.

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Acknowledgements

Our special thanks go to S. Swaminathan, C. Pickart, L. Hengst, A. Perrakis and members of the labs for stimulating discussions, T. Buesgen for her advice on GST-SUMO1 pull-downs, P. Celie for the CD experiments, H. Langedijk of Pepscan for the peptide synthesis, and A. Klanner, J. Vordemann and E. Stieger for excellent technical assistance. C. Pickart, N. Dantuma, A. Geerlof and R. Hay are gratefully acknowledged for providing reagents.

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Author notes

  1. Andrea Pichler and Puck Knipscheer: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Biochemistry I, University of Göttingen, Humboldt Allee 23, Göttingen, 37073, Germany

    Andrea Pichler & Frauke Melchior

  2. The Netherlands Cancer Institute, Plesmanlaan 121, Amsterdam, 1066 CX, The Netherlands

    Puck Knipscheer, Willem J van Dijk & Titia K Sixma

  3. Max-Planck Institute for Biochemistry, Am Klopferspitz 18, Martinsried, D-82152, Germany

    Andrea Pichler, Edith Oberhofer, Roman Körner, Stefan Jentsch & Frauke Melchior

  4. Department of Biochemistry and Molecular Biology, University of Southern Denmark, Campusvej 55, Odense M, Denmark

    Jesper Velgaard Olsen

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Correspondence to Andrea Pichler or Titia K Sixma.

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Supplementary information

Supplementary Fig. 1

Conventional SUMO sites in E2-25K structure. (PDF 1777 kb)

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Pichler, A., Knipscheer, P., Oberhofer, E. et al. SUMO modification of the ubiquitin-conjugating enzyme E2-25K. Nat Struct Mol Biol 12, 264–269 (2005). https://doi.org/10.1038/nsmb903

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