The structure of p53 tumour suppressor protein reveals the basis for its functional plasticity

p53 major tumour suppressor protein has presented a challenge for structural biology for two decades. The intact and complete p53 molecule has eluded previous attempts to obtain its structure, largely due to the intrinsic flexibility of the protein. Using ATP‐stabilised p53, we have employed cryoelectron microscopy and single particle analysis to solve the first three‐dimensional structure of the full‐length p53 tetramer (resolution 13.7 Å). The p53 molecule is a D2 tetramer, resembling a hollow skewed cube with node‐like vertices of two sizes. Four larger nodes accommodate central core domains, as was demonstrated by fitting of its X‐ray structure. The p53 monomers are connected via their juxtaposed N‐ and C‐termini within smaller N/C nodes to form dimers. The dimers form tetramers through the contacts between core nodes and N/C nodes. This structure revolutionises existing concepts of p53's molecular organisation and resolves conflicting data relating to its biochemical properties. This architecture of p53 in toto suggests novel mechanisms for structural plasticity, which enables the protein to bind variably spaced DNA target sequences, essential for p53 transactivation and tumour suppressor functions.

[1]  M. Heel,et al.  Exact filters for general geometry three dimensional reconstruction , 1986 .

[2]  W. O. Saxton,et al.  Principles of organization in S layers. , 1986, Journal of molecular biology.

[3]  M. Heel,et al.  Angular reconstitution: a posteriori assignment of projection directions for 3D reconstruction. , 1987 .

[4]  M. Radermacher,et al.  Three-dimensional reconstruction of single particles from random and nonrandom tilt series. , 1988, Journal of electron microscopy technique.

[5]  J. Milner,et al.  Cotranslation of activated mutant p53 with wild type drives the wild-type p53 protein into the mutant conformation , 1991, Cell.

[6]  K. Kinzler,et al.  Identification of p53 as a sequence-specific DNA-binding protein , 1991, Science.

[7]  D. Lane,et al.  Regulation of the specific DNA binding function of p53 , 1992, Cell.

[8]  K. Kinzler,et al.  Definition of a consensus binding site for p53 , 1992, Nature Genetics.

[9]  M. Remm,et al.  A C-terminal alpha-helix plus basic region motif is the major structural determinant of p53 tetramerization. , 1992, Oncogene.

[10]  S. Fields,et al.  Use of the two-hybrid system to identify the domain of p53 involved in oligomerization. , 1993, Oncogene.

[11]  M. Tarunina,et al.  Human p53 binds DNA as a protein homodimer but monomeric variants retain full transcription transactivation activity. , 1993, Oncogene.

[12]  P. Friedman,et al.  The p53 protein is an unusually shaped tetramer that binds directly to DNA. , 1993, Proceedings of the National Academy of Sciences of the United States of America.

[13]  P. Jeffrey,et al.  Crystal structure of a p53 tumor suppressor-DNA complex: understanding tumorigenic mutations. , 1994, Science.

[14]  J. E. Stenger,et al.  p53 oligomerization and DNA looping are linked with transcriptional activation. , 1994, The EMBO journal.

[15]  K. Kinzler,et al.  p53 tagged sites from human genomic DNA. , 1994, Human molecular genetics.

[16]  D. Lane,et al.  Allosteric activation of latent p53 tetramers , 1994, Current Biology.

[17]  J. E. Stenger,et al.  p53 domains: structure, oligomerization, and transformation , 1994, Molecular and cellular biology.

[18]  A. Levine,et al.  p53 and its 14 kDa C-terminal domain recognize primary DNA damage in the form of insertion/deletion mismatches , 1995, Cell.

[19]  Two domains of p53 interact with the TATA-binding protein, and the adenovirus 13S E1A protein disrupts the association, relieving p53-mediated transcriptional repression. , 1995, Molecular and cellular biology.

[20]  P. Tegtmeyer,et al.  Interaction of p53 with its consensus DNA-binding site , 1995, Molecular and cellular biology.

[21]  G. Marius Clore,et al.  Refined solution structure of the oligomerization domain of the tumour suppressor p53 , 1995, Nature Structural Biology.

[22]  L. Szekely,et al.  p53 binds single-stranded DNA ends through the C-terminal domain and internal DNA segments via the middle domain. , 1995, Nucleic acids research.

[23]  A. Levine,et al.  The carboxyl-terminal domain of the p53 protein regulates sequence-specific DNA binding through its nonspecific nucleic acid-binding activity. , 1995, Proceedings of the National Academy of Sciences of the United States of America.

[24]  N. Pavletich,et al.  Crystal structure of the tetramerization domain of the p53 tumor suppressor at 1.7 angstroms , 1995, Science.

[25]  A. Levine,et al.  Structure of the MDM2 Oncoprotein Bound to the p53 Tumor Suppressor Transactivation Domain , 1996, Science.

[26]  N. Pavletich,et al.  Structure of the p53 Tumor Suppressor Bound to the Ankyrin and SH3 Domains of 53BP2 , 1996, Science.

[27]  M van Heel,et al.  A new generation of the IMAGIC image processing system. , 1996, Journal of structural biology.

[28]  C. Prives,et al.  p53: puzzle and paradigm. , 1996, Genes & development.

[29]  Y. L. Lin,et al.  Dissection of Functional Domains of the Human DNA Replication Protein Complex Replication Protein A* , 1996, The Journal of Biological Chemistry.

[30]  P. Tegtmeyer,et al.  Reciprocal interference between the sequence-specific core and nonspecific C-terminal DNA binding domains of p53: implications for regulation , 1997, Molecular and cellular biology.

[31]  F. Ponchel,et al.  Induced N‐ and C‐terminal cleavage of p53: a core fragment of p53, generated by interaction with damaged DNA, promotes cleavage of the N‐terminus of full‐length p53, whereas ssDNA induces C‐terminal cleavage of p53 , 1997, The EMBO journal.

[32]  D. Lane,et al.  The N terminus of the murine p53 tumour suppressor is an independent regulatory domain affecting activation and thermostability. , 1998, Journal of molecular biology.

[33]  T. Halazonetis,et al.  Identification of an additional negative regulatory region for p53 sequence-specific DNA binding. , 1998, Proceedings of the National Academy of Sciences of the United States of America.

[34]  P. Tegtmeyer,et al.  Synergistic transcriptional activation of the MCK promoter by p53: tetramers link separated DNA response elements by DNA looping , 1998, Oncogene.

[35]  S. Halford,et al.  DNA looping by the SfiI restriction endonuclease , 1998 .

[36]  S. Halford,et al.  DNA looping by the Sfi I restriction endonuclease. , 1998, Journal of molecular biology.

[37]  C Urbanke,et al.  The Cfr10I restriction enzyme is functional as a tetramer. , 1999, Journal of molecular biology.

[38]  K. Wiman,et al.  Reactivation of Mutant p53 through Interaction of a C-Terminal Peptide with the Core Domain , 1999, Molecular and Cellular Biology.

[39]  Joseph R. Milner,et al.  An ATP/ADP-Dependent Molecular Switch Regulates the Stability of p53-DNA Complexes , 1999, Molecular and Cellular Biology.

[40]  R. Huber,et al.  Structure of the tetrameric restriction endonuclease NgoMIV in complex with cleaved DNA , 2000, Nature Structural Biology.

[41]  M. Heel,et al.  Single-particle electron cryo-microscopy: towards atomic resolution , 2000, Quarterly Reviews of Biophysics.

[42]  C. Prives,et al.  The N Terminus of p53 Regulates Its Dissociation from DNA* , 2000, The Journal of Biological Chemistry.

[43]  A. Levine,et al.  Surfing the p53 network , 2000, Nature.

[44]  J. Levine,et al.  Surfing the p53 network , 2000, Nature.

[45]  L. Kay,et al.  Latent and active p53 are identical in conformation , 2001, Nature Structural Biology.

[46]  Y. Jiao,et al.  Dynamic interactions of p53 with DNA in solution by time-lapse atomic force microscopy. , 2001, Journal of molecular biology.

[47]  J. Espinosa,et al.  Transcriptional regulation by p53 through intrinsic DNA/chromatin binding and site-directed cofactor recruitment. , 2001, Molecular cell.

[48]  R. Marmorstein,et al.  Crystal Structure of the Mouse p53 Core DNA-binding Domain at 2.7 Å Resolution* , 2001, The Journal of Biological Chemistry.

[49]  C. Prives,et al.  The C-terminus of p53: the more you learn the less you know , 2001, Nature Structural Biology.

[50]  T. Ekström,et al.  p53 latency--out of the blind alley. , 2002, Trends in biochemical sciences.

[51]  Jef D. Boeke,et al.  Structure of a Sir2 enzyme bound to an acetylated p53 peptide. , 2002, Molecular cell.

[52]  J. Lepault,et al.  Electronic Reprint Biological Crystallography on the Fitting of Model Electron Densities into Em Reconstructions: a Reciprocal-space Formulation , 2022 .

[53]  C. Prives,et al.  Efficient Specific DNA Binding by p53 Requires both Its Central and C-Terminal Domains as Revealed by Studies with High-Mobility Group 1 Protein , 2002, Molecular and Cellular Biology.

[54]  L. Serpell,et al.  Crystal structure of human 53BP1 BRCT domains bound to p53 tumour suppressor , 2002, The EMBO journal.

[55]  W. Deppert,et al.  The complex interactions of p53 with target DNA: we learn as we go. , 2003, Biochemistry and cell biology = Biochimie et biologie cellulaire.

[56]  P. Jeffrey,et al.  Structural differences in the DNA binding domains of human p53 and its C. elegans ortholog Cep-1. , 2004, Structure.

[57]  C. Prives,et al.  p53 linear diffusion along DNA requires its C terminus. , 2004, Molecular cell.

[58]  P. Jeffrey,et al.  Structural Differences in the DNA Binding Domains of Human p53 and its C. elegans Ortholog Cep-1: Structure of C. elegans Cep-1 , 2004 .

[59]  D. Scharre.  The Complex Interaction of Cognitive Issues , 2004 .

[60]  C. Harris,et al.  p53: traffic cop at the crossroads of DNA repair and recombination , 2005, Nature Reviews Molecular Cell Biology.

[61]  P. Hainaut,et al.  25 years of p53 research , 2005 .

[62]  C. Arrowsmith,et al.  Single-stranded DNA mimicry in the p53 transactivation domain interaction with replication protein A , 2005, Proceedings of the National Academy of Sciences of the United States of America.

[63]  Antonina Andreeva,et al.  Core domain interactions in full-length p53 in solution , 2006, Proceedings of the National Academy of Sciences of the United States of America.

[64]  Ronen Marmorstein,et al.  Structure of the p53 Core Domain Dimer Bound to DNA*♦ , 2006, Journal of Biological Chemistry.

[65]  M. Kitayner,et al.  Structural basis of DNA recognition by p53 tetramers. , 2006, Molecular cell.