PDBsum entry 3fea

Cell cycle

PDB id: 3fea

Contents

Protein chain

83 a.a.

Ligands

ACE-PHE-MET-AIB-PM3-6CW-GLU-1AC-LEU-NH2 ×2

MPD

Waters ×82

References listed in PDB file

Key reference

Title

Crystal structures of human mdmx (hdmx) in complex with p53 peptide analogues reveal surprising conformational changes.

Authors

J.Kallen, A.Goepfert, A.Blechschmidt, A.Izaac, M.Geiser, G.Tavares, P.Ramage, P.Furet, K.Masuya, J.Lisztwan.

Ref.

J Biol Chem, 2009, 284, 8812-8821. [DOI no: 10.1074/jbc.M809096200]

PubMed id

19153082

Abstract

p53 tumor suppressor activity is negatively regulated through binding to the oncogenic proteins Hdm2 and HdmX. The p53 residues Leu(26), Trp(23), and Phe(19) are crucial to mediate these interactions. Inhibiting p53 binding to both Hdm2 and HdmX should be a promising clinical approach to reactivate p53 in the cancer setting, but previous studies have suggested that the discovery of dual Hdm2/HdmX inhibitors will be difficult. We have determined the crystal structures at 1.3 A of the N-terminal domain of HdmX bound to two p53 peptidomimetics without and with a 6-chlorine substituent on the indole (which binds in the same subpocket as Trp(23) of p53). The latter compound is the most potent peptide-based antagonist of the p53-Hdm2 interaction yet to be described. The x-ray structures revealed surprising conformational changes of the binding cleft of HdmX, including an "open conformation" of Tyr(99) and unexpected "cross-talk" between the Trp and Leu pockets. Notably, the 6-chloro p53 peptidomimetic bound with high affinity to both HdmX and Hdm2 (K(d) values of 36 and 7 nm, respectively). Our results suggest that the development of potent dual inhibitors for HdmX and Hdm2 should be feasible. They also reveal possible conformational states of HdmX, which should lead to a better prediction of its interactions with potential biological partners.

Figure 1.
Overall crystal structure of human HdmX in complex with the peptide-analogue Ac-Phe-Met-Aib-Pmp-Trp-Glu-Ac[3]c-Leu-NH[2] (compound 1). A, van der Waals surface representation showing the complex between HdmX (carbons in yellow, nitrogens in blue, oxygens in red, and sulfurs in brown) and the p53 peptide analogue compound 1 (ball-and-stick-model, carbons in cyan). Selected water molecules and hydrogen bonds are shown in white. The pockets into which Phe^19, Trp^23, and Leu^26 of p53 bind are indicated. Compound 1 makes several water-mediated interactions with HdmX and two direct hydrogen bonds (with CO-Met^53 and OE1-Gln^71). The phosphonate of the Pmp residue does not make direct interactions with HdmX. The most important differences with Hdm2 are seen for the Leu pocket and the bottom of the Trp pocket (Fig. 3), whereas the Phe pockets are similar. B, chemical structures of compound 1 (Ac-Phe-Met-Aib-Pmp-Trp-Glu-Ac[3]c-Leu-NH[2]) and the derivative compound 2, which has a 6-chlorine substituent on the indole ring. The orientation of the chemical structure drawing is adapted to resemble A and C, i.e. with the C terminus on the left and the N terminus on the right. C, compound 1 fitted into the 2F[o]-F[c] electron density map. Figs. 1, A and C, 2, and 3 were generated with PyMOL (39).

Figure 3.
HdmX and Hdm2 have important differences in the Leu and Trp pockets. A, superposition of the complexes HdmX-compound 1 (color coding is as in Figs. 1A and 2, i.e. HdmX with carbons are in yellow, compound 1 with carbons are in cyan) and Hdm2-optimized p53 peptide (PDB entry code 1T4F; Hdm2 and ligand with carbons in white) zoomed in on the Leu and Trp pockets. Amino acid residues that differ in identity between HdmX and Hdm2 have two labels (upper label from Hdm2), otherwise only one label (numbering for HdmX). The differences Pro^95 versus His^96, Met^53 versus Leu^54, and Leu^98 versus Ile^99 for HdmX versus Hdm2 modify the shape of the Leu pocket. In particular, the presence and position of CB-Pro^95 for HdmX leads to a different position of the Leu side chain from the ligand. B, superposition of the complexes HdmX-compound 2 (color coding as in Fig. 2, i.e. HdmX with carbons in brown, compound 2 with carbons in magenta) and Hdm2-compound 2 (PDB entry code 2GV2; Hdm2 and ligand with carbons in white), zoomed in on the Leu and Trp pockets. The presence of a 6-chlorine substituent at the bottom of the Trp pocket leads to dramatic side chain movements of Leu^98, Tyr^99, and Pro^95 for HdmX, where as Hdm2 shows practically no changes (cf. Figs. 2 and 3A). The Leu side chain of compound 2 can now adopt a very similar position for the HdmX and Hdm2 complexes (because the Leu pocket has been widened for HdmX), in contrast to the situation with a 6-H substituent in the Trp pocket (Fig. 3A). C, same superposition as in Fig. 3B but zoomed in on the Trp pocket. The differences Leu^85 versus Phe^86 and Leu^98 versus Ile^99 for HdmX versus Hdm2 modify the shape of the Trp pocket. In particular, the bottom of the Trp pocket is not yet completely filled by the 6-chlorine for HdmX. The important van der Waals interactions made by the 6-chlorine with Phe^86 for Hdm2 are only partially substituted by hydrophobic interactions with Leu^98 for HdmX. The diagram is programmed for stereo viewing.

The above figures are reprinted by permission from the ASBMB: J Biol Chem (2009, 284, 8812-8821) copyright 2009.