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.

Architectural and mechanistic insights into an EHD ATPase involved in membrane remodelling

This article has been updated


The ability to actively remodel membranes in response to nucleotide hydrolysis has largely been attributed to GTPases of the dynamin superfamily, and these have been extensively studied1. Eps15 homology (EH)-domain-containing proteins (EHDs/RME-1/pincher) comprise a less-well-characterized class of highly conserved eukaryotic ATPases implicated in clathrin-independent endocytosis2, and recycling from endosomes3,4. Here we show that EHDs share many common features with the dynamin superfamily, such as a low affinity for nucleotides, the ability to tubulate liposomes in vitro, oligomerization around lipid tubules in ring-like structures and stimulated nucleotide hydrolysis in response to lipid binding. We present the structure of EHD2, bound to a non-hydrolysable ATP analogue, and provide evidence consistent with a role for EHDs in nucleotide-dependent membrane remodelling in vivo. The nucleotide-binding domain is involved in dimerization, which creates a highly curved membrane-binding region in the dimer. Oligomerization of dimers occurs on another interface of the nucleotide-binding domain, and this allows us to model the EHD oligomer. We discuss the functional implications of the EHD2 structure for understanding membrane deformation.

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

Relevant articles

Open Access articles citing this article.

Access options

Rent or buy this article

Prices vary by article type



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

Figure 1: EHD2 shares common properties with the dynamin superfamily.
Figure 2: The structure of EHD2.
Figure 3: Membrane binding and the role of ATP hydrolysis.
Figure 4: The EHD2 oligomer.

Change history

  • 18 October 2007

    The AOP version of this paper contained the term 'Epsin homology (EH)-domain...'. This was corrected to 'Eps15 homology (EH)-domain...' in the 18 October issue


  1. Praefcke, G. J. & McMahon, H. T. The dynamin superfamily: universal membrane tubulation and fission molecules? Nature Rev. Mol. Cell Biol. 5, 133–147 (2004)

    Article  CAS  Google Scholar 

  2. Shao, Y. et al. Pincher, a pinocytic chaperone for nerve growth factor/TrkA signaling endosomes. J. Cell Biol. 157, 679–691 (2002)

    Article  CAS  Google Scholar 

  3. Grant, B. et al. Evidence that RME-1, a conserved C. elegans EH-domain protein, functions in endocytic recycling. Nature Cell Biol. 3, 573–579 (2001)

    Article  ADS  CAS  Google Scholar 

  4. Caplan, S. et al. A tubular EHD1-containing compartment involved in the recycling of major histocompatibility complex class I molecules to the plasma membrane. EMBO J. 21, 2557–2567 (2002)

    Article  CAS  Google Scholar 

  5. Blume, J. J., Halbach, A., Behrendt, D., Paulsson, M. & Plomann, M. EHD proteins are associated with tubular and vesicular compartments and interact with specific phospholipids. Exp. Cell Res. 313, 219–231 (2007)

    Article  CAS  Google Scholar 

  6. George, M. et al. Shared as well as distinct roles of EHD proteins revealed by biochemical and functional comparisons in mammalian cells and C. elegans . BMC Cell Biol. 8, 3 (2007)

    Article  Google Scholar 

  7. Jovic, M., Naslavsky, N., Rapaport, D., Horowitz, M. & Caplan, S. EHD1 regulates β1 integrin endosomal transport: effects on focal adhesions, cell spreading and migration. J. Cell Sci. 120, 802–814 (2007)

    Article  CAS  Google Scholar 

  8. Lin, S. X., Grant, B., Hirsh, D. & Maxfield, F. R. Rme-1 regulates the distribution and function of the endocytic recycling compartment in mammalian cells. Nature Cell Biol. 3, 567–572 (2001)

    Article  CAS  Google Scholar 

  9. Park, S. Y. et al. EHD2 interacts with the insulin-responsive glucose transporter (GLUT4) in rat adipocytes and may participate in insulin-induced GLUT4 recruitment. Biochemistry 43, 7552–7562 (2004)

    Article  CAS  Google Scholar 

  10. Rotem-Yehudar, R., Galperin, E. & Horowitz, M. Association of insulin-like growth factor 1 receptor with EHD1 and SNAP29. J. Biol. Chem. 276, 33054–33060 (2001)

    Article  CAS  Google Scholar 

  11. Valdez, G. et al. Pincher-mediated macroendocytosis underlies retrograde signaling by neurotrophin receptors. J. Neurosci. 25, 5236–5247 (2005)

    Article  CAS  Google Scholar 

  12. Braun, A. et al. EHD proteins associate with syndapin I and II and such interactions play a crucial role in endosomal recycling. Mol. Biol. Cell 16, 3642–3658 (2005)

    Article  CAS  Google Scholar 

  13. Lee, D. W. et al. ATP binding regulates oligomerization and endosome association of RME-1 family proteins. J. Biol. Chem. 280, 17213–17220 (2005)

    Article  CAS  Google Scholar 

  14. Guilherme, A. et al. EHD2 and the novel EH domain binding protein EHBP1 couple endocytosis to the actin cytoskeleton. J. Biol. Chem. 279, 10593–10605 (2004)

    Article  CAS  Google Scholar 

  15. Ghosh, A., Praefcke, G. J., Renault, L., Wittinghofer, A. & Herrmann, C. How guanylate-binding proteins achieve assembly-stimulated processive cleavage of GTP to GMP. Nature 440, 101–104 (2006)

    Article  ADS  CAS  Google Scholar 

  16. Reubold, T. F. et al. Crystal structure of the GTPase domain of rat dynamin 1. Proc. Natl Acad. Sci. USA 102, 13093–13098 (2005)

    Article  ADS  CAS  Google Scholar 

  17. Low, H. H. & Lowe, J. A bacterial dynamin-like protein. Nature 444, 766–769 (2006)

    Article  ADS  CAS  Google Scholar 

  18. de Beer, T., Carter, R. E., Lobel-Rice, K. E., Sorkin, A. & Overduin, M. Structure and Asn-Pro-Phe binding pocket of the Eps15 homology domain. Science 281, 1357–1360 (1998)

    Article  ADS  CAS  Google Scholar 

  19. de Beer, T. et al. Molecular mechanism of NPF recognition by EH domains. Nature Struct. Biol. 7, 1018–1022 (2000)

    Article  CAS  Google Scholar 

  20. Marks, B. et al. GTPase activity of dynamin and resulting conformation change are essential for endocytosis. Nature 410, 231–235 (2001)

    Article  ADS  CAS  Google Scholar 

  21. Martens, S., Kozlov, M. M. & McMahon, H. T. How synaptotagmin promotes membrane fusion. Science 316, 1205–1208 (2007)

    Article  ADS  CAS  Google Scholar 

  22. Zimmerberg, J. & Kozlov, M. M. How proteins produce cellular membrane curvature. Nature Rev. Mol. Cell Biol. 7, 9–19 (2006)

    Article  CAS  Google Scholar 

  23. Lenzen, C., Cool, R. H. & Wittinghofer, A. Analysis of intrinsic and CDC25-stimulated guanine nucleotide exchange of p21ras-nucleotide complexes by fluorescence measurements. Methods Enzymol. 255, 95–109 (1995)

    Article  CAS  Google Scholar 

  24. Van Duyne, G. D., Standaert, R. F., Karplus, P. A., Schreiber, S. L. & Clardy, J. Atomic structures of the human immunophilin FKBP-12 complexes with FK506 and rapamycin. J. Mol. Biol. 229, 105–124 (1993)

    Article  CAS  Google Scholar 

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

    Article  CAS  Google Scholar 

  26. Sheldrick, G. M. & Schneider, T. R. SHELXL: High-resolution refinement. Methods Enzymol. 277, 319–343 (1997)

    Article  CAS  Google Scholar 

  27. de la Fortelle, E. & Bricogne, G. in Methods in Enzymology (eds Carter, C. W. Jr & Sweet, R. M.). 472–494 (1997)

    Google Scholar 

  28. McRee, D. E. XtalView/Xfit—A versatile program for manipulating atomic coordinates and electron density. J. Struct. Biol. 125, 156–65 (1999)

    Article  CAS  Google Scholar 

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

    Article  CAS  Google Scholar 

  30. Laskowski, R. A., Macarthur, M. W., Moss, D. S. & Thornton, J. M. Procheck—a program to check the stereochemical quality of protein structures. J. Appl. Crystallogr. 26, 283–291 (1993)

    Article  CAS  Google Scholar 

  31. Kraulis, P. J. Molscript—a program to produce both detailed and schematic plots of protein structures. J. Appl. Crystallogr. 24, 946–950 (1991)

    Article  Google Scholar 

  32. Merritt, E. A. & Murphy, M. E. Raster3D Version 2.0. A program for photorealistic molecular graphics. Acta Crystallogr. D 50, 869–73 (1994)

    Article  CAS  Google Scholar 

  33. Landau, M. et al. ConSurf 2005: the projection of evolutionary conservation scores of residues on protein structures. Nucleic Acids Res. 33, W299–W302 (2005)

    Article  ADS  CAS  Google Scholar 

  34. Potterton, E., McNicholas, S., Krissinel, E., Cowtan, K. & Noble, M. The CCP4 molecular graphics project. Acta Crystallogr. D 58, 1955–1957 (2002)

    Article  Google Scholar 

  35. DeLano, W. L. The PyMOL Molecular Graphics System (DeLano Scientific, Palo Alto, California, USA, 2002)

    Google Scholar 

  36. Guex, N. & Peitsch, M. C. SWISS-MODEL and the Swiss-PdbViewer: An environment for comparative protein modeling. Electrophor. 18, 2714–2723 (1997)

    Article  CAS  Google Scholar 

  37. Collaborative Computational Project The CCP4 suite: programs for protein crystallography. Acta Crystallogr . D 50, 760 (1994)

    Google Scholar 

  38. Philo, J. S. A method for directly fitting the time derivative of sedimentation velocity data and an alternative algorithm for calculating sedimentation coefficient distribution functions. Analyt. Biochem. 279, 151–163 (2000)

    Article  CAS  Google Scholar 

  39. Philo, J. S. Improved methods for fitting sedimentation coefficient distributions derived by time-derivative techniques. Analyt. Biochem. 354, 238–246 (2006)

    Article  CAS  Google Scholar 

  40. Laue, T. M., Shah, B. D., Ridgeway, T. M. & Pelletier, S. L. in Analytical Ultracentrifugation in Biochemistry and Polymer Science (eds Harding, S. E., Rowe, A. J. & Horton, J. C.) 90–125 (Roy. Soc. of Chem., Cambridge, UK, 1992)

    Google Scholar 

Download references


Long-term fellowships supported O.D. (The International Human Frontier Science Program Organization), R.L. (Swedish Research Council) and S.M. (EMBO). We thank M. Plomann for providing the complementary DNAs for mammalian EHDs, and the ESRF beam staff in Grenoble for their support. The authors declare no competing financial interests.

The atomic coordinates of mouse EHD2 have been deposited in the Protein Data Bank (PDB) with the accession number 2QPT.

Author information

Authors and Affiliations


Corresponding authors

Correspondence to Oliver Daumke or Harvey T. McMahon.

Ethics declarations

Competing interests

Reprints and permissions information is available at The authors declare no competing financial interests.

Supplementary information

Supplementary Information 1

This file contains Supplementary Figures 1-12 with Legends, Supplementary Table 1 with the data collection statistics and additional references. (PDF 2225 kb)

Supplementary Information 2

The file contains Supplementary Video 1 which shows EHD2 wild-type was over-expressed in HeLa cells for 24 h and imaged by EPI-fluorescence for approximately 30 min. Some of the tubules and puncta are dynamic. (MOV 4707 kb)

Supplementary Information 3

The file contains Supplementary Video 2 which shows cropped area of EHD2 WT (Video 1) at higher resolution. (MOV 5319 kb)

Supplementary Information 4

The file contains Supplementary Video 3 which shows EHD2 T94A was over-expressed in HeLa cells for 24 h and imaged by EPI-fluorescence for approximately 30 min. There are only tubules, and these are mostly static. Cropped movies show a smaller area at higher resolution. (MOV 6652 kb)

Supplementary Information 5

The file contains Supplementary Video 4 which shows cropped area of EHD2 T94A (Video 2) at higher resolution. (MOV 7726 kb)

Supplementary Information 6

The file contains Supplementary Video 5 which shows EHD2 I157Q was over-expressed in HeLa cells for 24h and imaged by EPI-fluorescence for approximately 30min. No tubules can be found and the puncta are mostly motile. (MOV 9013 kb)

Supplementary Information 7

The file contains Supplementary Video 6 which shows cropped area of EHD2 I157Q (Video 4) at higher resolution. (MOV 6353 kb)

Supplementary Information 8

The file contains Supplementary Data with PDB coordinates of the proposed EHD2 oligomer. Four EHD2 dimers (in the absence of the EH domain) were aligned as described in Methods. All lipid interaction sites point towards the putative membrane interface. Molecules B and C which have been used for the initial alignment with GBP1 are related via a 2-fold axis and the nucleotides of these molecules are oriented in a head-to-head fashion. (ZIP 455 kb)

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Daumke, O., Lundmark, R., Vallis, Y. et al. Architectural and mechanistic insights into an EHD ATPase involved in membrane remodelling. Nature 449, 923–927 (2007).

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI:

This article is cited by


By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.


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