Modulation of p53 binding to MDM2: Computational studies reveal important roles of Tyr100

#07-01 Matrix, Singapore.
BMC Bioinformatics (Impact Factor: 2.58). 12/2009; 10 Suppl 15(Suppl 15):S6. DOI: 10.1186/1471-2105-10-S15-S6
Source: PubMed


The tumor suppressor protein p53 is regulated by the ubiquitin ligase MDM2 which down-regulates p53. In tumours with overexpressed MDM2, the p53-MDM2 interaction can be interrupted by a peptide or small molecule to stabilize p53 as a therapeutic strategy. Structural and biochemical/mutagenesis data show that p53 has 3 hydrophobic residues F19, W23 and L26 that embed into the ligand binding pocket of MDM2 which is highly plastic in nature and can modulate its size to accommodate a variety of ligands. This binding pocket is primarily dependent on the orientation of a particular residue, Y100. We have studied the role of the dynamics of Y100 in p53 recognition.
Molecular dynamics simulations show that the Y100 side chain can be in "open" or "closed" states with only the former enabling complex formation. When both p53 and MDM2 are in near native conformations, complex formation is rapid and is driven by the formation of a hydrogen bond between W23 of p53 and L54 of MDM2 or by the embedding of F19 of p53 into MDM2. The transition of Y100 from "closed" to "open" can increase the size of the binding site. Interconversions between these two states can be induced by the N-terminal region of MDM2 or by the conformations of the p53 peptides.
Molecular dynamics simulations have revealed how the binding of p53 to MDM2 is modulated by the conformational mobility of Y100 which is the gatekeeper residue in MDM2. The mobility of this residue can be modulated by the conformations of p53 and the Nterminal lid region of MDM2.

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    • "Key to this interaction is a triad of p53 amino acids that insert deeply into the MDM2 cleft: Phe19, Trp23, and Leu26 (designated as three subpockets on MDM2; Figure 1; Figure S1 available online). The MDM2-p53 recognition is a dynamic and multistage process that employs the bindinginduced folding of p53 (Kussie et al., 1996; Popowicz et al., 2011; Uesugi and Verdine, 1999; Lee et al., 2000), the rearrangement of the Leu26 subpocket of MDM2 by a twist of the Tyr100 ring from the ''closed'' to the ''open'' (anti)conformations (as defined by Popowicz et al., 2007, 2008; Kussie et al., 1996; Dastidar et al., 2009; Uhrinova et al., 2005), and the dissociation of a transient a-helical N-terminal ''lid'' segment of MDM2 (residues 19–23) from the proximity of the p53-complementary interface (McCoy et al., 2003; Showalter et al., 2008; Zhan et al., 2012). Until now, all structurally characterized low-molecular-weight inhibitors of the MDM2-p53 interaction targeted the same ''closed'' Tyr100 state and were incapable of reaching the N-terminal ''lid'' segment, an intrinsically disordered region of MDM2 (Popowicz et al., 2011; Shangary and Wang, 2009; Dö mling, 2008; Graves et al., 2012). "
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    ABSTRACT: Reactivation of p53 by release of the functional protein from its inhibition by MDM2 provides an efficient, nongenotoxic approach to a wide variety of cancers. We present the cocrystal structures of two complexes of MDM2 with inhibitors based on 6-chloroindole scaffolds. Both molecules bound to a distinct conformational state of MDM2 with nM-μM affinities. In contrast to other structurally characterized antagonists, which mimic three amino acids of p53 (Phe19, Trp23, and Leu26), the compounds induced an additional hydrophobic pocket on the MDM2 surface and unveiled a four-point binding mode. The enlarged interaction interface of the inhibitors resulted in extension of small molecules binding toward the “lid” segment of MDM2 (residues 19–23)—a nascent element that interferes with p53 binding. As supported by protein engineering and molecular dynamics studies, employing these unstable elements of MDM2 provides an efficient and yet unexplored alternative in development of MDM2-p53 association inhibitors.
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    • "Although MDM2 and MDMX proteins are highly homologous, they display different dynamic behavior. More recently, Dastidar et al. [88] emphasized the role of the dynamics of Tyr100, which was recognized as a putative gatekeeper in p53-MDM2 interactions by MD simulations. It appears that the transition of the Tyr100 side chain from “closed” to “open” enlarges the binding site, thus enhancing the binding of MDM2. "
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    ABSTRACT: As a tumor suppressor protein, p53 plays a crucial role in the cell cycle and in cancer prevention. Almost 50 percent of all human malignant tumors are closely related to a deletion or mutation in p53. The activity of p53 is inhibited by over-active celluar antagonists, especially by the over-expression of the negative regulators MDM2 and MDMX. Protein-protein interactions, or post-translational modifications of the C-terminal negative regulatory domain of p53, also regulate its tumor suppressor activity. Restoration of p53 function through peptide and small molecular inhibitors has become a promising strategy for novel anti-cancer drug design and development. Molecular dynamics simulations have been extensively applied to investigate the conformation changes of p53 induced by protein-protein interactions and protein-ligand interactions, including peptide and small molecular inhibitors. This review focuses on the latest MD simulation research, to provide an overview of the current understanding of interactions between p53 and its partners at an atomic level.
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    • "A general trend observed from the simulations was that the last two C-terminal residues of the peptides undergo a transition from an extended strand conformation to a helical conformation within a few nanoseconds (see Movies S1). This change in peptide conformation, coupled with the flipping of Y100 (MDM2) towards the ligand binding pocket, creates a more complementary fit for the peptide in the MDM2 binding pocket as has been reported previously [11], [13], [21], [22]. Without the rotation of the Y100 side chain, the C-terminal residues in the peptide would not make any specific contacts with MDM2. "
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    ABSTRACT: The complex between the proteins MDM2 and p53 is a promising drug target for cancer therapy. The residues 19-26 of p53 have been biochemically and structurally demonstrated to be a most critical region to maintain the association of MDM2 and p53. Variation of the amino acid sequence in this range obviously alters the binding affinity. Surprisingly, suitable substitutions contiguous to this region of the p53 peptides can yield tightly binding peptides. The peptide variants may differ by a single residue that vary little in their structural conformations and yet are characterized by large differences in their binding affinities. In this study a systematic analysis into the role of single C-terminal mutations of a 12 residue fragment of the p53 transactivation domain (TD) and an equivalent phage optimized peptide (12/1) were undertaken to elucidate their mechanistic and thermodynamic differences in interacting with the N-terminal of MDM2. The experimental results together with atomistically detailed dynamics simulations provide insight into the principles that govern peptide design protocols with regard to protein-protein interactions and peptidomimetic design.
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