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:

Molecular basis for N-terminal acetylation by the heterodimeric NatA complex

Nature Structural & Molecular Biology volume 20, pages 1098–1105 (2013)Cite this article

Subjects

Abstract

N-terminal acetylation is ubiquitous among eukaryotic proteins and controls a myriad of biological processes. Of the N-terminal acetyltransferases (NATs) that facilitate this cotranslational modification, the heterodimeric NatA complex has the most diversity for substrate selection and modifies the majority of all N-terminally acetylated proteins. Here, we report the X-ray crystal structure of the 100-kDa holo-NatA complex from Schizosaccharomyces pombe, in the absence and presence of a bisubstrate peptide-CoA–conjugate inhibitor, as well as the structure of the uncomplexed Naa10p catalytic subunit. The NatA-Naa15p auxiliary subunit contains 13 tetratricopeptide motifs and adopts a ring-like topology that wraps around the NatA-Naa10p subunit, an interaction that alters the Naa10p active site for substrate-specific acetylation. These studies have implications for understanding the mechanistic details of other NAT complexes and how regulatory subunits modulate the activity of the broader family of protein acetyltransferases.

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

Access options

Buy this article

  • Purchase on SpringerLink
  • Instant access to full article PDF
Buy now

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

Figure 1: Overall structure of the NatA complex bound to acetyl CoA.
Figure 2: Structure of the Naa10p monomer bound to acetyl CoA.
Figure 3: Inhibitor structures and IC50 curves.
Figure 4: Structure of the NatA complex bound to a bisubstrate inhibitor.
Figure 5: The active site of the NatA complex.

Similar content being viewed by others

Accession codes

Primary accessions

Protein Data Bank

Referenced accessions

Protein Data Bank

References

  1. Arnesen, T. et al. Proteomics analyses reveal the evolutionary conservation and divergence of N-terminal acetyltransferases from yeast and humans. Proc. Natl. Acad. Sci. USA 106, 8157–8162 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. Forte, G.M., Pool, M.R. & Stirling, C.J. N-terminal acetylation inhibits protein targeting to the endoplasmic reticulum. PLoS Biol. 9, e1001073 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Hwang, C.S., Shemorry, A. & Varshavsky, A. N-terminal acetylation of cellular proteins creates specific degradation signals. Science 327, 973–977 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Scott, D.C., Monda, J.K., Bennett, E.J., Harper, J.W. & Schulman, B.A. N-terminal acetylation acts as an avidity enhancer within an interconnected multiprotein complex. Science 334, 674–678 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Starheim, K.K., Gevaert, K. & Arnesen, T. Protein N-terminal acetyltransferases: when the start matters. Trends Biochem. Sci. 37, 152–161 (2012).

    Article  CAS  PubMed  Google Scholar 

  6. Yi, C.H. et al. Metabolic regulation of protein N-alpha-acetylation by Bcl-xL promotes cell survival. Cell 146, 607–620 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Arnesen, T. et al. Identification and characterization of the human ARD1-NATH protein acetyltransferase complex. Biochem. J. 386, 433–443 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Gautschi, M. et al. The yeast Nα-acetyltransferase NatA is quantitatively anchored to the ribosome and interacts with nascent polypeptides. Mol. Cell Biol. 23, 7403–7414 (2003).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Mullen, J.R. et al. Identification and characterization of genes and mutants for an N-terminal acetyltransferase from yeast. EMBO J. 8, 2067–2075 (1989).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Park, E.C. & Szostak, J.W. ARD1 and NAT1 proteins form a complex that has N-terminal acetyltransferase activity. EMBO J. 11, 2087–2093 (1992).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Polevoda, B., Brown, S., Cardillo, T.S., Rigby, S. & Sherman, F. Yeast Nα-terminal acetyltransferases are associated with ribosomes. J. Cell Biochem. 103, 492–508 (2008).

    Article  CAS  PubMed  Google Scholar 

  12. Starheim, K.K. et al. Identification of the human Nα-acetyltransferase complex B (hNatB): a complex important for cell-cycle progression. Biochem. J. 415, 325–331 (2008).

    Article  CAS  PubMed  Google Scholar 

  13. Starheim, K.K. et al. Knockdown of human Nα-terminal acetyltransferase complex C leads to p53-dependent apoptosis and aberrant human Arl8b localization. Mol. Cell Biol. 29, 3569–3581 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. Arnesen, T., Thompson, P.R., Varhaug, J.E. & Lillehaug, J.R. The protein acetyltransferase ARD1: a novel cancer drug target? Curr. Cancer Drug Targets 8, 545–553 (2008).

    Article  CAS  PubMed  Google Scholar 

  15. Arnesen, T. et al. Induction of apoptosis in human cells by RNAi-mediated knockdown of hARD1 and NATH, components of the protein N-α-acetyltransferase complex. Oncogene 25, 4350–4360 (2006).

    Article  CAS  PubMed  Google Scholar 

  16. Lim, J.H., Park, J.W. & Chun, Y.S. Human arrest defective 1 acetylates and activates β-catenin, promoting lung cancer cell proliferation. Cancer Res. 66, 10677–10682 (2006).

    Article  CAS  PubMed  Google Scholar 

  17. Ren, T. et al. Generation of novel monoclonal antibodies and their application for detecting ARD1 expression in colorectal cancer. Cancer Lett. 264, 83–92 (2008).

    Article  CAS  PubMed  Google Scholar 

  18. Kalvik, T.V. & Arnesen, T. Protein N-terminal acetyltransferases in cancer. Oncogene 32, 269–276 (2013).

    Article  CAS  PubMed  Google Scholar 

  19. Yu, M. et al. Correlation of expression of human arrest-defective-1 (hARD1) protein with breast cancer. Cancer Invest. 27, 978–983 (2009).

    Article  CAS  PubMed  Google Scholar 

  20. Fluge, Ø., Bruland, O., Akslen, L.A., Varhaug, J.E. & Lillehaug, J.R. NATH, a novel gene overexpressed in papillary thyroid carcinomas. Oncogene 21, 5056–5068 (2002).

    Article  CAS  PubMed  Google Scholar 

  21. Yu, M. et al. Immunohistochemical analysis of human arrest-defective-1 expressed in cancers in vivo. Oncol. Rep. 21, 909–915 (2009).

    CAS  PubMed  Google Scholar 

  22. Polevoda, B. & Sherman, F. Composition and function of the eukaryotic N-terminal acetyltransferase subunits. Biochem. Biophys. Res. Commun. 308, 1–11 (2003).

    Article  CAS  PubMed  Google Scholar 

  23. Polevoda, B. & Sherman, F. N-terminal acetyltransferases and sequence requirements for N-terminal acetylation of eukaryotic proteins. J. Mol. Biol. 325, 595–622 (2003).

    Article  CAS  PubMed  Google Scholar 

  24. Starheim, K.K., Gromyko, D., Velde, R., Varhaug, J.E. & Arnesen, T. Composition and biological significance of the human Nα-terminal acetyltransferases. BMC Proc. 3 (suppl. 6), S3 (2009).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  25. Polevoda, B., Norbeck, J., Takakura, H., Blomberg, A. & Sherman, F. Identification and specificities of N-terminal acetyltransferases from Saccharomyces cerevisiae. EMBO J. 18, 6155–6168 (1999).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Van Damme, P. et al. Proteome-derived peptide libraries allow detailed analysis of the substrate specificities of Nα-acetyltransferases and point to hNaa10p as the post-translational actin Nα-acetyltransferase. Mol. Cell Proteomics 10, M110.004580 (2011).

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  27. Van Damme, P. et al. N-terminal acetylome analyses and functional insights of the N-terminal acetyltransferase NatB. Proc. Natl. Acad. Sci. USA 109, 12449–12454 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Sutton, A. et al. Sas4 and Sas5 are required for the histone acetyltransferase activity of Sas2 in the SAS complex. J. Biol. Chem. 278, 16887–16892 (2003).

    Article  CAS  PubMed  Google Scholar 

  29. Iizuka, M. & Stillman, B. Histone acetyltransferase HBO1 interacts with the ORC1 subunit of the human initiator protein. J. Biol. Chem. 274, 23027–23034 (1999).

    Article  CAS  PubMed  Google Scholar 

  30. Parthun, M.R., Widom, J. & Gottschling, D.E. The major cytoplasmic histone acetyltransferase in yeast: links to chromatin replication and histone metabolism. Cell 87, 85–94 (1996).

    Article  CAS  PubMed  Google Scholar 

  31. Polevoda, B., Hoskins, J. & Sherman, F. Properties of Nat4, an Nα-acetyltransferase of Saccharomyces cerevisiae that modifies N termini of histones H2A and H4. Mol. Cell Biol. 29, 2913–2924 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Hole, K. et al. The human N-alpha-acetyltransferase 40 (hNaa40p/hNatD) is conserved from yeast and N-terminally acetylates histones H2A and H4. PLoS ONE 6, e24713 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Van Damme, P. et al. NatF contributes to an evolutionary shift in protein N-terminal acetylation and is important for normal chromosome segregation. PLoS Genet. 7, e1002169 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Song, O.K., Wang, X., Waterborg, J.H. & Sternglanz, R. An Nα-acetyltransferase responsible for acetylation of the N-terminal residues of histones H4 and H2A. J. Biol. Chem. 278, 38109–38112 (2003).

    Article  CAS  PubMed  Google Scholar 

  35. Evjenth, R. et al. Human Naa50p (Nat5/San) displays both protein Nα- and Nå-acetyltransferase activity. J. Biol. Chem. 284, 31122–31129 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. Liszczak, G., Arnesen, T. & Marmorstein, R. Structure of a ternary Naa50p (NAT5/SAN) N-terminal acetyltransferase complex reveals the molecular basis for substrate-specific acetylation. J. Biol. Chem. 286, 37002–37010 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  37. Blatch, G.L. & Lassle, M. The tetratricopeptide repeat: a structural motif mediating protein-protein interactions. Bioessays 21, 932–939 (1999).

    Article  CAS  PubMed  Google Scholar 

  38. Arnesen, T. et al. The chaperone-like protein HYPK acts together with NatA in cotranslational N-terminal acetylation and prevention of Huntingtin aggregation. Mol. Cell Biol. 30, 1898–1909 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  39. Cingolani, G., Petosa, C., Weis, K. & Muller, C.W. Structure of importin-β bound to the IBB domain of importin-α. Nature 399, 221–229 (1999).

    Article  CAS  PubMed  Google Scholar 

  40. Vetting, M.W. et al. Structure and functions of the GNAT superfamily of acetyltransferases. Arch. Biochem. Biophys. 433, 212–226 (2005).

    Article  CAS  PubMed  Google Scholar 

  41. Tanner, K.G. et al. Catalytic mechanism and function of invariant glutamic acid 173 from the histone acetyltransferase GCN5 transcriptional coactivator. J. Biol. Chem. 274, 18157–18160 (1999).

    Article  CAS  PubMed  Google Scholar 

  42. Berndsen, C.E., Albaugh, B.N., Tan, S. & Denu, J.M. Catalytic mechanism of a MYST family histone acetyltransferase. Biochemistry 46, 623–629 (2007).

    Article  CAS  PubMed  Google Scholar 

  43. Evjenth, R.H. et al. Human protein N-terminal acetyltransferase hNaa50p (hNAT5/hSAN) follows ordered sequential catalytic mechanism: combined kinetic and NMR study. J. Biol. Chem. 287, 10081–10088 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. Braman, J., Papworth, C. & Greener, A. Site-directed mutagenesis using double-stranded plasmid DNA templates. Methods Mol. Biol. 57, 31–44 (1996).

    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. McCoy, A.J. et al. Phaser crystallographic software. J. Appl. Crystallogr. 40, 658–674 (2007).

    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  CAS  Google Scholar 

  48. Langer, G., Cohen, S.X., Lamzin, V.S. & Perrakis, A. Automated macromolecular model building for X-ray crystallography using ARP/wARP version 7. Nat. Protoc. 3, 1171–1179 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  49. 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 

  50. Painter, J. & Merritt, E.A. A molecular viewer for the analysis of TLS rigid-body motion in macromolecules. Acta Crystallogr. D Biol. Crystallogr. 61, 465–471 (2005).

    Article  PubMed  CAS  Google Scholar 

  51. Painter, J. & Merritt, E.A. Optimal description of a protein structure in terms of multiple groups undergoing TLS motion. Acta Crystallogr. D Biol. Crystallogr. 62, 439–450 (2006).

    Article  PubMed  CAS  Google Scholar 

  52. Terwilliger, T.C. & Berendzen, J. Automated MAD and MIR structure solution. Acta Crystallogr. D Biol. Crystallogr. 55, 849–861 (1999).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  53. Terwilliger, T.C. Maximum-likelihood density modification. Acta Crystallogr. D Biol. Crystallogr. 56, 965–972 (2000).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

This work was supported by US National Institutes of Health (NIH) grant GM060293 (R.M.) and NIH training grant GM071339 (G.L.). We acknowledge the use of the Wistar Proteomics Core facility for the work reported here, which is supported in part by NIH grant CA010815. T.A. was supported by the Research Council of Norway and the Norwegian Cancer Society. We also acknowledge Marmorstein laboratory members and E. Skordalakes for helpful discussions.

Author information

Authors and Affiliations

  1. Program in Gene Expression and Regulation, Wistar Institute, Philadelphia, Pennsylvania, USA

    Glen Liszczak & Ronen Marmorstein

  2. Department of Chemistry, University of Pennsylvania, Philadelphia, Pennsylvania, USA

    Glen Liszczak, Jacob M Goldberg, E James Petersson & Ronen Marmorstein

  3. Department of Molecular Biology, University of Bergen, Bergen, Norway

    Håvard Foyn & Thomas Arnesen

  4. Department of Surgery, Haukeland University Hospital, Bergen, Norway

    Thomas Arnesen

Authors
  1. Glen Liszczak
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Jacob M Goldberg
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Håvard Foyn
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. E James Petersson
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Thomas Arnesen
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Ronen Marmorstein
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

G.L. performed all of the structural and biochemical experiments described in the manuscript, and J.M.G. carried out inhibitor synthesis. G.L. prepared manuscript figures, text and videos; H.F. carried out preliminary inhibition studies that led to experiments reported in the manuscript; R.M. designed and supervised experiments by G.L. and prepared manuscript text. T.A. supervised the experiments of H.F. and prepared manuscript text. E.J.P. supervised the experiments of J.M.G. All authors read and approved the submitted manuscript.

Corresponding author

Correspondence to Ronen Marmorstein.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–5 (PDF 12310 kb)

An overall view of the NatA complex structure

The view shown in Fig. 1A is shown rotating 360° on the y-axis followed by 360° on the x-axis. (AVI 17786 kb)

Global conformational shifts in Naa10p upon Naa15p binding

This video begins with the uncomplexed Naa10p and highlights all residues featured in Figures 1c,d. It first shows the position of these residues in the uncomplexed Naa10p and a morph shows the change in position of these residues upon Naa15p binding. (AVI 29209 kb)

Naa10p active site conformational changes upon Naa15p binding

This movie shows a morph that highlights the change in position of Naa10p residues Leu22, Glu24, Tyr26, Arg113 and Tyr139 upon Naa15p binding. The morph is shown twice, once with AcCoA bound to Naa10p and again with the bisubstrate inhibitor-bound Naa10p. (AVI 12648 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Liszczak, G., Goldberg, J., Foyn, H. et al. Molecular basis for N-terminal acetylation by the heterodimeric NatA complex. Nat Struct Mol Biol 20, 1098–1105 (2013). https://doi.org/10.1038/nsmb.2636

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nsmb.2636

This article is cited by

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