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.

Full-length p53 tetramer bound to DNA and its quaternary dynamics

Oncogene volume 36, pages 1451–1460 (2017)Cite this article

Subjects

Abstract

P53 is a major tumor suppressor that is mutated and inactivated in ~50% of all human cancers. Thus, reactivation of mutant p53 using small molecules has been a long sought-after anticancer therapeutic strategy. Full structural characterization of the full-length oligomeric p53 is challenging because of its complex architecture and multiple highly flexible regions. To explore p53 structural dynamics, here we developed a series of atomistic integrative models with available crystal structures of the full-length p53 (fl-p53) tetramer bound to three different DNA sequences: a p21 response element, a puma response element and a nonspecific DNA sequence. Explicitly solvated, all-atom molecular dynamics simulations of the three complexes (totaling nearly 1 μs of aggregate simulation time) yield final structures consistent with electron microscopy maps and, for the first time, show the direct interactions of the p53 C-terminal with DNA. Through a collective principal component analysis, we identify sequence-dependent differential quaternary binding modes of the p53 tetramer interfacing with DNA. Additionally, L1 loop dynamics of fl-p53 in the presence of DNA is revealed, and druggable pockets of p53 are identified via solvent mapping to aid future drug discovery studies.

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

from$1.95

to$39.95

Learn more

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

Figure 1
Figure 2
Figure 3
Figure 4

References

  1. Vogelstein B, Lane D, Levine AJ . Surfing the p53 network. Nature 2000; 408: 307–310.

    Article  CAS  PubMed  Google Scholar 

  2. Vousden KH, Lu X . Live or let die: the cell's response to p53. Nat Rev Cancer 2002; 2: 594–604.

    Article  CAS  PubMed  Google Scholar 

  3. Green DR, Kroemer G . Cytoplasmic functions of the tumour suppressor p53. Nature 2009; 458: 1127–1130.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Li T, Kon N, Jiang L, Tan M, Ludwig T, Zhao Y et al. Tumor suppression in the absence of p53-mediated cell-cycle arrest, apoptosis, and senescence. Cell 2012; 149: 1269–1283.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Valente LJ, Gray DH, Michalak EM, Pinon-Hofbauer J, Egle A, Scott CL et al. P53 efficiently suppresses tumor development in the complete absence of its cell-cycle inhibitory and proapoptotic effectors p21, Puma, and Noxa. Cell Rep 2013; 3: 1339–1345.

    Article  CAS  PubMed  Google Scholar 

  6. Wassman CD, Baronio R, Demir O, Wallentine BD, Chen CK, Hall LV et al. Computational identification of a transiently open L1/S3 pocket for reactivation of mutant p53. Nat Commun 2013; 4: 1407.

    Article  PubMed  Google Scholar 

  7. Joerger AC, Fersht AR . The tumor suppressor p53: from structures to drug discovery. Cold Spring Harb Perspect Biol 2010; 2: a000919.

    Article  PubMed  PubMed Central  Google Scholar 

  8. Lambert JM, Gorzov P, Veprintsev DB, Soderqvist M, Segerback D, Bergman J et al. PRIMA-1 reactivates mutant p53 by covalent binding to the core domain. Cancer Cell 2009; 15: 376–388.

    Article  CAS  PubMed  Google Scholar 

  9. Liu X, Wilcken R, Joerger AC, Chuckowree IS, Amin J, Spencer J et al. Small molecule induced reactivation of mutant p53 in cancer cells. Nucleic Acids Res 2013; 41: 6034–6044.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  10. Yu X, Vazquez A, Levine AJ, Carpizo DR . Allele-specific p53 mutant reactivation. Cancer Cell 2012; 21: 614–625.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Joerger AC, Bauer MR, Wilcken R, Baud MG, Harbrecht H, Exner TE et al. Exploiting transient protein states for the design of small-molecule stabilizers of mutant p53. Structure 2015; 23: 2246–2255.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Ventura A, Kirsch DG, McLaughlin ME, Tuveson DA, Grimm J, Lintault L et al. Restoration of p53 function leads to tumour regression in vivo. Nature 2007; 445: 661–665.

    Article  CAS  PubMed  Google Scholar 

  13. Martins CP, Brown-Swigart L, Evan GI . Modeling the therapeutic efficacy of p53 restoration in tumors. Cell 2006; 127: 1323–1334.

    Article  CAS  PubMed  Google Scholar 

  14. Wiman KG . Restoration of wild-type p53 function in human tumors: strategies for efficient cancer therapy. Adv Cancer Res 2007; 97: 321–338.

    Article  CAS  PubMed  Google Scholar 

  15. Tidow H, Melero R, Mylonas E, Freund SM, Grossmann JG, Carazo JM et al. Quaternary structures of tumor suppressor p53 and a specific p53 DNA complex. Proc Natl Acad Sci USA 2007; 104: 12324–12329.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Fink AL . Natively unfolded proteins. Curr Opin Struct Biol 2005; 15: 35–41.

    Article  CAS  PubMed  Google Scholar 

  17. Hupp TR . Regulation of p53 protein function through alterations in protein-folding pathways. Cell Mol Life Sci 1999; 55: 88–95.

    Article  CAS  PubMed  Google Scholar 

  18. Emamzadah S, Tropia L, Halazonetis TD . Crystal structure of a multidomain human p53 tetramer bound to the natural CDKN1A (p21) p53-response element. Mol Cancer Res 2011; 9: 1493–1499.

    Article  CAS  PubMed  Google Scholar 

  19. Emamzadah S, Tropia L, Vincenti I, Falquet B, Halazonetis TD . Reversal of the DNA-binding-induced loop L1 conformational switch in an engineered human p53 protein. J Mol Biol 2014; 426: 936–944.

    Article  CAS  PubMed  Google Scholar 

  20. Petty TJ, Emamzadah S, Costantino L, Petkova I, Stavridi ES, Saven JG et al. An induced fit mechanism regulates p53 DNA binding kinetics to confer sequence specificity. EMBO J 2011; 30: 2167–2176.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Kitayner M, Rozenberg H, Kessler N, Rabinovich D, Shaulov L, Haran TE et al. Structural basis of DNA recognition by p53 tetramers. Mol Cell 2006; 22: 741–753.

    Article  CAS  PubMed  Google Scholar 

  22. Weinberg RL, Veprintsev DB, Bycroft M, Fersht AR . Comparative binding of p53 to its promoter and DNA recognition elements. J Mol Biol 2005; 348: 589–596.

    Article  CAS  PubMed  Google Scholar 

  23. Ahn J, Prives C . The C-terminus of p53: the more you learn the less you know. Nat Struct Biol 2001; 8: 730–732.

    Article  CAS  PubMed  Google Scholar 

  24. Hupp TR, Meek DW, Midgley CA, Lane DP . Regulation of the specific DNA binding function of p53. Cell 1992; 71: 875–886.

    Article  CAS  PubMed  Google Scholar 

  25. McKinney K, Mattia M, Gottifredi V, Prives C . P53 linear diffusion along DNA requires its C terminus. Mol Cell. 2004; 16: 413–424.

    Article  CAS  PubMed  Google Scholar 

  26. Tafvizi A, Huang F, Fersht AR, Mirny LA, van Oijen AM . A single-molecule characterization of p53 search on DNA. Proc Natl Acad Sci USA 2011; 108: 563–568.

    Article  CAS  PubMed  Google Scholar 

  27. Lukman S, Lane DP, Verma CS . Mapping the structural and dynamical features of multiple p53 DNA binding domains: insights into loop 1 intrinsic dynamics. PLoS One 2013; 8: e80221.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Melero R, Rajagopalan S, Lazaro M, Joerger AC, Brandt T, Veprintsev DB et al. Electron microscopy studies on the quaternary structure of p53 reveal different binding modes for p53 tetramers in complex with DNA. Proc Natl Acad Sci USA 2011; 108: 557–562.

    Article  CAS  PubMed  Google Scholar 

  29. Nagaich AK, Zhurkin VB, Durell SR, Jernigan RL, Appella E, Harrington RE . P53-induced DNA bending and twisting: p53 tetramer binds on the outer side of a DNA loop and increases DNA twisting. Proc Natl Acad Sci USA 1999; 96: 1875–1880.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Balagurumoorthy P, Sakamoto H, Lewis MS, Zambrano N, Clore GM, Gronenborn AM et al. Four p53 DNA-binding domain peptides bind natural p53-response elements and bend the DNA. Proc Natl Acad Sci USA 1995; 92: 8591–8595.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Weinberg RL, Veprintsev DB, Fersht AR . Cooperative binding of tetrameric p53 to DNA. J Mol Biol 2004; 341: 1145–1159.

    Article  CAS  PubMed  Google Scholar 

  32. Lavery R, Moakher M, Maddocks JH, Petkeviciute D, Zakrzewska K . Conformational analysis of nucleic acids revisited: Curves+. Nucleic Acids Res 2009; 37: 5917–5929.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. D'Abramo M, Besker N, Desideri A, Levine AJ, Melino G, Chillemi G . The p53 tetramer shows an induced-fit interaction of the C-terminal domain with the DNA-binding domain. Oncogene 2016; 35: 3272–3281.

    Article  CAS  PubMed  Google Scholar 

  34. Arlt C, Ihling CH, Sinz A . Structure of full-length p53 tumor suppressor probed by chemical cross-linking and mass spectrometry. Proteomics 2015; 15: 2746–2755.

    Article  CAS  PubMed  Google Scholar 

  35. Friedler A, Veprintsev DB, Freund SM, von Glos KI, Fersht AR . Modulation of binding of DNA to the C-terminal domain of p53 by acetylation. Structure 2005; 13: 629–636.

    Article  CAS  PubMed  Google Scholar 

  36. Viadiu H, Aggarwal AK . Structure of BamHI bound to nonspecific DNA: a model for DNA sliding. Mol Cell 2000; 5: 889–895.

    Article  CAS  PubMed  Google Scholar 

  37. Leith JS, Tafvizi A, Huang F, Uspal WE, Doyle PS, Fersht AR et al. Sequence-dependent sliding kinetics of p53. Proc Natl Acad Sci USA 2012; 109: 16552–16557.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Molecular Operating Environment (MOE) H3A 2R7. 2013.08 ed. Chemical Computing Group Inc.: Montreal, QC, Canada, 2016.

  39. Humphrey W, Dalke A, Schulten K . VMD: visual molecular dynamics. J Mol Graph 1996; 14: 33–38 27–28.

    Article  CAS  PubMed  Google Scholar 

  40. Jacobs SA, Harp JM, Devarakonda S, Kim Y, Rastinejad F, Khorasanizadeh S . The active site of the SET domain is constructed on a knot. Nat Struct Biol 2002; 9: 833–838.

    CAS  PubMed  Google Scholar 

  41. Maestro, version 10.4 Schrodinger, LLC: New York, NY, USA, 2015.

  42. Natan E, Baloglu C, Pagel K, Freund SM, Morgner N, Robinson CV et al. Interaction of the p53 DNA-binding domain with its n-terminal extension modulates the stability of the p53 tetramer. J Mol Biol 2011; 409: 358–368.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  43. Lim KW, Amrane S, Bouaziz S, Xu W, Mu Y, Patel DJ et al. Structure of the human telomere in K+ solution: a stable basket-type G-quadruplex with only two G-tetrad layers. J Am Chem Soc 2009; 131: 4301–4309.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. Bochkareva E, Kaustov L, Ayed A, Yi GS, Lu Y, Pineda-Lucena A et al. Single-stranded DNA mimicry in the p53 transactivation domain interaction with replication protein A. Proc Natl Acad Sci USA 2005; 102: 15412–15417.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Rustandi RR, Baldisseri DM, Weber DJ . Structure of the negative regulatory domain of p53 bound to S100B(betabeta). Nat Struct Biol 2000; 7: 570–574.

    Article  CAS  PubMed  Google Scholar 

  46. Lowe ED, Tews I, Cheng KY, Brown NR, Gul S, Noble ME et al. Specificity determinants of recruitment peptides bound to phospho-CDK2/cyclin A. Biochemistry 2002; 41: 15625–15634.

    Article  CAS  PubMed  Google Scholar 

  47. Ma B, Pan Y, Zheng J, Levine AJ, Nussinov R . Sequence analysis of p53 response-elements suggests multiple binding modes of the p53 tetramer to DNA targets. Nucleic Acids Res 2007; 35: 2986–3001.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  48. Jorgensen WL, Chandrasekhar J, D. MJ, Impey RW, Klein ML . Comparison of simple potential functions for simulating liquid water. J Chem Phys 1983; 79: 926–935.

    Article  CAS  Google Scholar 

  49. Case DA, Berryman JT, Betz RM, Cerutti DS, Cheatham TE III, Darden TA et al. AMBER 2015. University of California: San Francisco, CA, USA, 2015.

    Google Scholar 

  50. Pang YP . Novel zinc protein molecular dynamics simulations: steps toward antiangiogenesis for cancer treatment. J Mol Model 1999; 5: 196–202.

    Article  CAS  Google Scholar 

  51. Maier JA, Martinez C, Kasavajhala K, Wickstrom L, Hauser KE, Simmerling C . ff14SB: improving the accuracy of protein side chain and backbone parameters from ff99SB. J Chem Theory Comput 2015; 11: 3696–3713.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  52. Darden T, Perera L, Li L, Pedersen L . New tricks for modelers from the crystallography toolkit: the particle mesh Ewald algorithm and its use in nucleic acid simulations. Structure 1999; 7: R55–R60.

    Article  CAS  PubMed  Google Scholar 

  53. Phillips JC, Braun R, Wang W, Gumbart J, Tajkhorshid E, Villa E et al. Scalable molecular dynamics with NAMD. J Comput Chem 2005; 26: 1781–1802.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  54. Wickham H . Ggplot2 Elegant Graphics for Data Analysis. Springer: New York, NY, USA, 2009.

    Google Scholar 

  55. Janert PK . Gnuplot in Action: Understanding Data with Graphs. Manning: Greenwich, CT, USA, 2010.

  56. Sanner MF . Python: a programming language for software integration and development. J Mol Graph Model 1999; 17: 57–61.

    CAS  PubMed  Google Scholar 

  57. Durrant JD, Votapka L, Sorensen J, Amaro RE . POVME 2.0: an enhanced tool for determining pocket shape and volume characteristics. J Chem Theory Comput 2014; 10: 5047–5056.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  58. Altintas I, Berkley C, Jaeger E, Jones M, Ludascher B, Mock S . Kepler: an extensible system for design and execution of scientific workflows. 16th International Conference on Scientific and Statistical Database Management, 2004: Proceedings 2004, pp 423–424.

  59. R Core Team. A Language and Environment for Statistical Computing. R Foundation for Statistical Computing: Vienna, Austria, 2015.

  60. Ngan CH, Bohnuud T, Mottarella SE, Beglov D, Villar EA, Hall DR et al. FTMAP: extended protein mapping with user-selected probe molecules. Nucleic Acids Res 2012; 40: W271–W275.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  61. Trabuco LG, Villa E, Mitra K, Frank J, Schulten K . Flexible fitting of atomic structures into electron microscopy maps using molecular dynamics. Structure 2008; 16: 673–683.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  62. Pettersen EF, Goddard TD, Huang CC, Couch GS, Greenblatt DM, Meng EC et al. UCSF Chimera—a visualization system for exploratory research and analysis. J Comput Chem 2004; 25: 1605–1612.

    Article  CAS  PubMed  Google Scholar 

  63. Votapka LW, Czapla L, Zhenirovskyy M, Amaro RE . DelEnsembleElec: computing ensemble-averaged electrostatics using DelPhi. Commun Comput Phys 2013; 13: 256–268.

    Article  Google Scholar 

Download references

Acknowledgements

We thank Mikel Valle for kindly providing the three-dimensional cryo-EM maps in 2011 Melero et al. article, and Steffen Lindert, Rob Swift, Clarisse Ricci, Lane Votapka, Jacob Durrant and Elizabeth Villa for helpful discussions. This work was supported by the University of California Cancer Research Coordinating Committee award and NIH Innovator award DP2-007237. Computing time on Stampede and Comet supercomputers was provided through the NSF XSEDE supercomputer resources grant RAC CHE060073N (to REA). Additional support from the National Biomedical Computation Resource NIH P41 GM103426 is acknowledged.

Author contributions

ÖD, PUI and REA designed research; ÖD designed the model; PUI performed the simulations; ÖD and PUI analyzed data; ÖD, PUI, and REA wrote the paper.

Author information

Author notes

  1. Ö Demir and P U Ieong: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Chemistry and Biochemistry, University of California, San Diego, La Jolla, CA, USA

    Ö Demir, P U Ieong & R E Amaro

  2. National Biomedical Computation Resource, University of California, San Diego, La Jolla, CA, USA

    P U Ieong & R E Amaro

Authors
  1. Ö Demir
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. P U Ieong
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. R E Amaro
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to R E Amaro.

Ethics declarations

Competing interests

REA is a co-founder of Actavalon Inc., a company developing p53 reactivation compounds for anticancer therapeutics.

Additional information

Supplementary Information accompanies this paper on the Oncogene website

Supplementary information

Rights and permissions

Reprints and Permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Demir, Ö., Ieong, P. & Amaro, R. Full-length p53 tetramer bound to DNA and its quaternary dynamics. Oncogene 36, 1451–1460 (2017). https://doi.org/10.1038/onc.2016.321

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/onc.2016.321

This article is cited by

Search

Advanced search

Quick links