- P53 is a tumor suppressor protein that acts as a transcription factor, forming a homotetramer. - It is activated by various stresses like DNA damage or hypoxia to arrest the cell cycle or induce apoptosis. - P53 levels are kept low through a negative feedback loop, as it induces transcription of its negative regulator MDM2. - The protein contains distinct domains for transcription activation, DNA binding, oligomerization and nuclear localization that allow it to perform its tumor suppressor functions.

1. STRUCTURE AND FUNCTION OF
P53
PROTEIN
AHMAD ALI
DEPARTMENT OF MOLECULAR DIAGNOSTIC AND BIOTECHNOLOGY
THE UNIVERSITY OF CAMERINO, ITALY

2. OUTLINES
• INTRODUCTION
• LOCATION OF THE GENE
• STRUCTURE OF THE P53
• P53 DOMAINS AND ITS FUNCTION
• CONTROL OF P53
• REFERENCES

3. INTRODUCTION
• The p53 tumor suppressor protein is a classic gatekeeper of cellular fate. It is a tumor
suppressor gene and the most frequent site of genetic alterations found in human
cancer.
• The p53 protein is a transcription factor that regulates the expression of a wide
variety of genes involved in cell cycle arrest and apoptosis in response to Genotoxic
or cellular stress.
• The protein was discovered because it bound to the large T antigen in SV40 infected
cells and was therefore co immuno-precipitated with antibodies generated against the
viral protein.

4. P53 RESPONSE TO:
• P53 restricts tumor development by serving as a sensor of cellular stress, responding to diverse
signals, including…..
• DNA damage, hypoxia, oncogene expression, limiting the propagation of cells under these
adverse conditions.
• Conditions of low-level stress:
Alternatively, under conditions of low-level stress, p53 elicits protective, pro-survival
responses, such as temporary cell-cycle arrest, DNA repair and antioxidant protein production, to
maintain genome integrity and viability in cells that sustain limited.
• When an experiment was performed in cells with wild-type p53 alleles, the p53 protein
disappeared with a half-life of only 20 minutes. Its located in ER, mitochondria and nucleus.

5. p53-activating signals and p53’s downstream effects

7. LOCATION OF GENE
• In humans, the TP53 gene is located on the short arm of chromosome 17. The MW of the protein
is 53 kDa.
• The gene spans 20 kb, with a very long first intron of 10 kb.
• The coding sequence contains five regions showing a high degree of conservation in vertebrates,
predominantly in exons 2, 5, 6, 7 and 8.
Figure adapted from Proceedings of the
National Academy of Sciences of United
states of America.

8. STRUCTURE OF P53
• Wild-type p53 protein contains 393 amino acids and is composed of several structural and
functional domains. It’s a homotetramer.
1. N-terminus containing an amino-terminal domain (residues 1- 42).
2. A proline-rich region with multiple copies of the PXXP sequence (residues 61-94, where X is
any amino acid).
3. A central core domain (residues 102-292), and a C terminal region (residues 301-393).
4. It contains an oligomerization domain (residues 324-355).
5. A strongly basic carboxyl terminal regulatory domain (residues 363-393).

10. FUNCTION OF DOMAINS
• N-TERMINAL DOMAIN:
The n-terminus of p53 comprises two Transcriptional
activation domains (Tads), TAD1 and TAD2 . These
Domains can independently enhance Transcription of
p53 target genes.
• They are required for transactivation activity and
interact with various transcription factors including
acetyltransferases and MDM2.
Structure of the MDM2 oncoprotein
bound to the p53 tumor suppressor
transactivation domain

11. PROLINE DOMAIN :
• P53 is a tumor suppressor gene which can activate or repress transcription, as well as induce
apoptosis. The human p53 Proline-rich domain localized between amino acids 64 and 92 has
Been reported to be necessary for efficient growth suppression.
• The proline-rich region plays a role in p53 stability regulated by MDM2, wherein p53 becomes
more susceptible to degradation by MDM2 if this region is deleted.
Central DNA binding domain:
• The central core of this protein is made up primarily of the DNA binding domain required for
sequence-specific DNA binding (the consensus sequence contains two copies of the10-bp motif
5’- PuPuPuC(A/T)-(T/A)GPyPyPy-3’, separated by 0-13 bp).
• Most cancer associated p53 mutations are missense mutations in this domain and incapacitate
DNA binding, illuminating the key importance of DNA binding for p53- mediated tumor
suppression.

12. Location of p53 point mutations

13. • Tetramerization domain:
• It is a part of C terminal domain of p53 protein present from 324-355. It is
well established that the p53 forms tetramers via an oligomerization domain.
• Tetramerization appears to be required for efficient transactivation in vivo
and for p53-mediated suppression of growth of carcinoma cell lines.
• Tetramerization domain ties the four chains together. A long flexible region
in each chain then connects to the DNA-binding domain.

14. PDB: 1olg- High-resolution structure of
the oligomerization domain of p53 by
multidimensional NMR Clore GM,
Omichinski JG, Sakaguchi

15. • C-TERMINAL BASIC DOMAIN:
• P53 contains a basic, lysine-rich domain which also functions as a negative
regulatory domain. According to the allosteric model, In which c-terminal tail of
p53 was considered as a negative Regulator and may regulate the ability of its core
DNA binding Domain to lock the DNA binding domain as an latent Conformation.
• An interaction between the C-terminal domain and another region in a p53
tetramer locks the tetramer in a DNA binding incompetent state. The activation of
p53 in cultured mammalian cells has been correlated with phosphorylation and
proteolytic removal of the C-terminal domain.
• These modifications are thought to activate p53 by causing a conformational
change of the protein, regulated by an allosteric effect.

16. • NUCLEAR LOCALIZATION SIGNALING:
Nuclear localization signaling (NLS) domain, residues 316-325 have been
identified in the c-terminal region. Mutagenesis of the most n-terminal signal
(NLS1, amino acids 316 ± 325) induces the synthesis of a totally
cytoplasmic p53 protein, while alteration Of the NLS2 (amino acids 369 ±
375) and NLS3 (amino acids 379 ± 384) leads to both cytoplasmic and
nuclear localization.

17. CONTROL OF P53 LEVELS BY MDM2
• After p53 concentrations increase in response to certain physiologic signals, the p53 tetramers
bind to the promoters of target genes whose transcription they induce.
• Among the induced genes is mdm2.
• Once synthesized, Mdm2 molecules bind to the p53 protein subunits and trigger their
ubiquitylation and export to the cytoplasm, where they are degraded in proteasomes.
• This negative-feedback loop ensures that p53 levels eventually sink back to a low level and, in
undisturbed cells, helps to keep p53 levels very low.

19. SUMMARY
• P53 is a transcription factor, acting as a homotetramer.
• Hyper-proliferative stress (e.g. oncogenic signaling, hypoxia) activates p53 via ARF
• Transcribed constitutively, but has a very short half life
• Ubiquitinated by the E3 ligase MDM2

20. REFERENCES
• Http://asia.Ensembl.Org/homo_sapiens/location/view?R=17:7565097 7590868
• Kussie, P.H., Gorina, S., Marechal, V., Elenbaas, B., Moreau, J..Levine, A.J., Pavletich, N.P
‘Structure of the MDM2 oncoprotein bound to the p53 tumor suppressor transactivation
domain’.
• Venot C, Maratrat M, Dureuil C, Conseiller E, Bracco L, Debussche L. The requirement for the
p53 proline-rich functional domain for mediation of apoptosis is correlated with specific PIG3
gene transactivation and with transcriptional repression. The EMBO Journal. 1998;17(16):4668-
4679. doi:10.1093/emboj/17.16.4668.
• Bai, Ling and Wei-Guo Zhu. “p 53 : Structure , Function and Therapeutic Applications.” (2006)
• May, Pierre, and Evelyne May. "Twenty years of p53 research: structural and functional aspects
of the p53 protein." Oncogene 18.53 (1999): 7621.
• Brady, Colleen A., and Laura D. Attardi. "p53 at a glance." J Cell Sci 123.15 (2010): 2527-2532.

21. THANK YOU SO MUCH !
WE MATURE WITH DAMAGE, NOT
YEAR’S………………