Molecular Basis for the Inhibition of p53 by Mdmx

Popowicz, Grzegorz M.; Czarna, Anna; Rothweiler, U.; Szwagierczak, Aleksandra; Krajewski, Marcin; Weber, Lutz; Holak, Tad A.

doi:10.4161/cc.6.19.4740

Cited by 124 publications

(184 citation statements)

References 46 publications

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“…11,52,53 While the detailed mechanism of regulation of p53, MDM2 and MDM4 is complex, the N-terminal region of MDM4 is known to modulate the levels of p53 too. A recent structural study of the N-terminal domain of MDM4 complexed with the transactivation region of p53 19 has identified some structural differences, mainly in the orientation of residues in one part of the p53-binding cleft, which may underlie the differential interactions between p53 and MDM4 (compared to p53 and MDM2). 18 Electrostatic maps of MDM4 (Fig.…”

Section: Resultsmentioning

confidence: 99%

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Modulation of the p53-MDM2 Interaction by Phosphorylation of Thr18: A Computational Study

Lee

Srinivasan²,

Coomber³

et al. 2007

Cell Cycle

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The transcription factor p53 is under negative regulation by the ubiquitin ligase MDM2 and its close homologue MDM4. In the bound complex between MDM2 and p53, the transactivation domain of p53 adopts an amphipathic helical conformation which optimizes the spatial organization of three key hydrophobic residues (Phe19, Trp23, Leu26) for maximum interactions. The interaction with MDM2 is known to be abrogated by phosphorylation of Ser/Thr residues in the MDM2 N-terminal domain and in the p53 transactivation domain. In the latter, phosphorylation of Thr18 has been attributed to destabilize a key hbond between Thr18 and Asp21. This interaction has been thought to be critical for the formation of the helical conformation of the p53 transactivation domain. Molecular dynamics simulations of the p53 transactivation domain suggest that phosphorylation of either Thr18 or Ser20 does not disrupt its helical structure but does result in reduced affinities for MDM2. While interactions between the Thr18 and Asp21 are indeed broken due to charge-charge repulsions, the peptide has enough inherent flexibility to form alternate patterns of hbonds, resulting in the maintenance of helicity. Electrostatics of MDM2 reveal local anionic patches in the region where Thr18 docks. These suggest that repulsions will arise because the MDM2 surface will force the p53 to bind in a manner that will place the negatively charged phosphorylated Thr18 near this anionic region. A similar, albeit somewhat attenuated pattern of electrostatic modulations, is seen for a model of MDM4 that has been built. Mutants of MDM2 and MDM4 have been designed to attenuate this anionicity and have been computationally demonstrated to enhance the binding of the phosphorylated peptides.

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Section: Resultsmentioning

confidence: 99%

“…18 A structural study of a "humanized" zebrafish MDM4 where the p53-binding site residues of zebrafish MDM4 have been mutated to those of the human form has been reported. 19 Their main findings were that a part of the p53-binding cleft is altered in its conformation compared to that seen in the structure of MDM2.…”

Section: Introductionmentioning

confidence: 99%

Modulation of the p53-MDM2 Interaction by Phosphorylation of Thr18: A Computational Study

Lee

Srinivasan²,

Coomber³

et al. 2007

Cell Cycle

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“…However, because of low solubility and relatively weak activity of pDI, this work focused only on PMI, which is highly soluble in aqueous solution and of higher affinity for MDM2 and MDMX than pDI. Two p53 peptides, (15)(16)(17)(18)(19)(20)(21)(22)(23)(24)(25)(26)(27)(28)(29) p53 (SQETFSDLWKLLPEN) and (17)(18)(19)(20)(21)(22)(23)(24)(25)(26)(27)(28) p53 (ETFSDLWKLLPE), were used for comparison.…”

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confidence: 99%

Structural basis for high-affinity peptide inhibition of p53 interactions with MDM2 and MDMX

Pazgier

Liu

Zou

et al. 2009

Proc. Natl. Acad. Sci. U.S.A.

354

514

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The oncoproteins MDM2 and MDMX negatively regulate the activity and stability of the tumor suppressor protein p53-a cellular process initiated by MDM2 and/or MDMX binding to the Nterminal transactivation domain of p53. MDM2 and MDMX in many tumors confer p53 inactivation and tumor survival, and are important molecular targets for anticancer therapy. We screened a duodecimal peptide phage library against site-specifically biotinylated p53-binding domains of human MDM2 and MDMX chemically synthesized via native chemical ligation, and identified several peptide inhibitors of the p53-MDM2/MDMX interactions. The most potent inhibitor (TSFAEYWNLLSP), termed PMI, bound to MDM2 and MDMX at low nanomolar affinities-approximately 2 orders of magnitude stronger than the wild-type p53 peptide of the same length (ETFSDLWKLLPE). We solved the crystal structures of synthetic MDM2 and MDMX, both in complex with PMI, at 1.6 Å resolution. Comparative structural analysis identified an extensive, tightened intramolecular H-bonding network in bound PMI that contributed to its conformational stability, thus enhanced binding to the 2 oncogenic proteins. Importantly, the C-terminal residue Pro of PMI induced formation of a hydrophobic cleft in MDMX previously unseen in the structures of p53-bound MDM2 or MDMX. Our findings deciphered the structural basis for highaffinity peptide inhibition of p53 interactions with MDM2 and MDMX, shedding new light on structure-based rational design of different classes of p53 activators for potential therapeutic use.

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“…Recent structural studies using humanized zebra fish MDMX and human MDMX in complex with the p53 N-terminal peptide revealed extensive similarity between the p53-binding domains of MDM2 and MDMX in overall folding and the shapes of their p53-binding pockets. However, a few sequence differences result in a smaller hydrophobic cleft in MDMX that prevents efficient binding by Nutlin (28,29). Future development of inhibitors against MDMX can be facilitated by identification of a high affinity artificial ligand that target MDMX and/or MDM2.…”

mentioning

confidence: 99%

Structure-based Design of High Affinity Peptides Inhibiting the Interaction of p53 with MDM2 and MDMX

Phan

Kasprzak

et al. 2010

Journal of Biological Chemistry

146

157

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MDM2 and MDMX function as key regulators of p53 by binding to its N terminus, inhibiting its transcriptional activity, and promoting degradation. MDM2 and MDMX overexpression or hyperactivation directly contributes to the loss of p53 function during the development of nearly 50% of human cancers. Recent studies showed that disrupting p53-MDM2 and p53-MDMX interactions can lead to robust activation of p53 but also revealed a need to develop novel dual specific or MDMXspecific inhibitors. Using phage display we identified a 12-residue peptide (pDI) with inhibitory activity against MDM2 and MDMX. The co-crystal structures of the pDI and a single mutant derivative (pDI6W) liganded with the N-terminal domains of human MDMX and MDM2 served as the basis for the design of 11 distinct pDI-derivative peptides that were tested for inhibitory potential. The best derivative (termed pDIQ) contained four amino acid substitutions and exhibited a 5-fold increase in potency over the parent peptide against both MDM2 (IC 50 ‫؍‬ 8 nM) and MDMX (IC 50 ‫؍‬ 110 nM). Further structural studies revealed key molecular features enabling the high affinity binding of the pDIQ to these proteins. These include large conformational changes of the pDIQ to reach into a hydrophobic site unique to MDMX. The findings suggest new strategies toward the rational design of small molecule inhibitors efficiently targeting MDMX.The p53 tumor suppressor is a potent inducer of cell cycle arrest, apoptosis, cellular senescence, and innate immunity. It is activated in response to oncogenic transformation, extrinsic stress, and viral infection to protect higher organisms from cancer (1-3). p53 also facilitates maternal reproduction through induction of the growth factor leukemia inhibitory factor (LIF) that promotes embryo implantation (4). p53 activity is kept at minimal levels in unstressed cells by interactions with MDM2 and MDMX. MDM2 is an ubiquitin E3 ligase for p53 and an important regulator of p53 stability by forming a negative feedback loop (5, 6). The MDM2 homolog MDMX also binds to p53 and inhibits p53-dependent transcription (7). Loss of MDM2 or MDMX leads to embryonic lethality (8 -10). Therefore, the expression of MDM2 and the expression of MDMX are both necessary for regulation of p53 during normal development.Genetic or functional inactivation of p53 is an obligatory step during cancer development. In human tumors that retain wild type p53, amplification of MDM2 or MDMX serves as an alternative mechanism of p53 inactivation in a subset of tumors (11,12). Furthermore, MDM2 activity is controlled by the tumor suppressor ARF (alternative reading frame) encoded by the INK4a locus (2). Deletion/epigenetic silencing of ARF occur in most tumors expressing wild type p53, resulting in hyperactive MDM2 and lack of p53 response to oncogenic stress in malignant tumors (13,14). ARF has also been shown to promote MDMX degradation by MDM2 (15). Loss of ARF expression may result in MDMX stabilization that further inactivates p53. Therefore, MDM2 and MDMX are d...

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Molecular Basis for the Inhibition of p53 by Mdmx

Cited by 124 publications

References 46 publications

Modulation of the p53-MDM2 Interaction by Phosphorylation of Thr18: A Computational Study

Modulation of the p53-MDM2 Interaction by Phosphorylation of Thr18: A Computational Study

Structural basis for high-affinity peptide inhibition of p53 interactions with MDM2 and MDMX

Structure-based Design of High Affinity Peptides Inhibiting the Interaction of p53 with MDM2 and MDMX

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