PDBsum entry 2v8x

Translation

PDB id: 2v8x

Contents

Protein chains

181 a.a.

14 a.a.

Ligands

MGQ ×2

Waters ×343

References listed in PDB file

Key reference

Title

Crystallographic and mass spectrometric characterisation of eif4e with n7-Alkylated cap derivatives.

Authors

C.J.Brown, I.Mcnae, P.M.Fischer, M.D.Walkinshaw.

Ref.

J Mol Biol, 2007, 372, 7. [DOI no: 10.1016/j.jmb.2007.06.033]

PubMed id

17631896

Abstract

Structural complexes of the eukaryotic translation initiation factor 4E (eIF4E) with a series of N(7)-alkylated guanosine derivative mRNA cap analogue structures have been characterised. Mass spectrometry was used to determine apparent gas-phase equilibrium dissociation constants (K(d)) values of 0.15 microM, 13.6 microM, and 55.7 microM for eIF4E with 7-methyl-GTP (m(7)GTP), GTP, and GMP, respectively. For tight and specific binding to the eIF4E mononucleotide binding site, there seems to be a clear requirement for guanosine derivatives to possess both the delocalised positive charge of the N(7)-methylated guanine system and at least one phosphate group. We show that the N(7)-benzylated monophosphates 7-benzyl-GMP (Bn(7)GMP) and 7-(p-fluorobenzyl)-GMP (FBn(7)GMP) bind eIF4E substantially more tightly than non-N(7)-alkylated guanosine derivatives (K(d) values of 7.0 microM and 2.0 microM, respectively). The eIF4E complex crystal structures with Bn(7)GMP and FBn(7)GMP show that additional favourable contacts of the benzyl groups with eIF4E contribute binding energy that compensates for loss of the beta and gamma-phosphates. The N(7)-benzyl groups pack into a hydrophobic pocket behind the two tryptophan side-chains that are involved in the cation-pi stacking interaction between the cap and the eIF4E mononucleotide binding site. This pocket is formed by an induced fit in which one of the tryptophan residues involved in cap binding flips through 180 degrees relative to structures with N(7)-methylated cap derivatives. This and other observations made here will be useful in the design of new families of eIF4E inhibitors, which may have potential therapeutic applications in cancer.

Figure 1.
Figure 1. Chemical structures of the mRNA cap and some cap derivatives. The N^7-substituted GMP cap analogues were synthesised from GMP and an appropriate benzyl halide. Experimental details are given as Supplementary Data.

Figure 3.
Figure 3. Electron density and electrostatic surface of three ligands m^7GTP, Bn^7GMP and FBn^7GMP bound to eIF4E. (a), (c) and (e) The F[o]–F[c] electron density (2.5σ) for the ligands m^7GTP, Bn^7GMP and FBn^7GMP, respectively, in blue, with the green mesh showing the 2 F[o]–F[c] map for eIF4E (1.2σ) and the structured waters. Incorporation of the bulky benzyl group in Bn^7GMP and FBn^7GMP causes the W102 side-chain to flip 180°. The m^7G moiety of the cap analogue forms the same hydrogen bonds (shown in yellow) with the E103 side-chain and the amide backbone of W102. (b), (d) and (f) The electrostatic potential of the eIF4E surface. The phosphate-binding region has a distinctive positive charge whilst the pocket in which the m^7G moiety intercalates is more hydrophobic in nature. The m^7G moiety forms a parallel stack with W102 and W56 and interacts with the two ring systems via cation–π interactions.

The above figures are reprinted by permission from Elsevier: J Mol Biol (2007, 372, 7-0) copyright 2007.