Messenger RNA 5' Cap-Binding Protein

Haichuan Gu '27 and Kunqi Shi '28

Contents:

I. Introduction

The eukaryotic messenger RNA 5' capping protein (eIF4E) is an RNA-binding protein involved in translation initiation. eIF4E recognizes the 5' cap structure of the mRNA, which consists of a 7-methylated guanosine connected with the first nucleotide of the mRNA via a 5'-5' triphosphate bridge. eIF4E is a component of the eIF4F complex, which consists of eIF4A, eIF4E, and eIF4G.

Upon eIF4E binding to the 5' cap, eIF4F and eIF4B unwinds the secondary structure of the 5' untranslated region of the mRNA and exposes the start codon to the ribosome. Because it is the least abundant translation initiation factor, eIF4E is considered to be the rate-limiting factor in the recruitment of the ribosome to the start codon.  

II. General Structure

eIF4E is a monomer of 45,210 molecular weight, composing of one polypeptide chain 217 amino acids of length. While the N-terminal portion (the first 35 residues) is divergent among different organisms and and is therefore omitted from the structure, the C-terminal portion (the last 182 residues) is highly conserved and required for cap recognition, suppression via binding to 4E-binding proteins, and stimulation of cap-dependent translation. Hightlighted in blue is the entire eIF4E molecule, and the other one is a duplicate. The protein adopts the shape of a "cupped hand", with 8 beta strands arranged into a curved, antiparallel beta sheet, 3 long alpha helices lying parallel to the strand direction and on the convex side of the beta sheet, and 1 additional short alpha helix on the concave side of the sheet. Meanwhile, the cap-binding site consists of the beta sheet, the short alpha helix , a loop between strands S1 and S2, and another loop between strands S3 and S4.  

III. RNA 5' Cap Binding

Once eIF4E engages the mRNA 5’ cap, it forms a compact recognition pocket that clamps the cap analog 7-methyl-GDP through four cooperative sets of interactions. First, the alkylated base 7-methyl-guanine is held in a “sandwich-like” π–π stacking arrangement between two conserved tryptophan residues, Trp-56 and Trp-102 These aromatic rings stack above and below the methylated guanine and provide the core of base recognition. Second, hydrogen bonds and van der Waals contacts between eIF4E and the 7-methyl-guanine further stabilize this interaction. In particular, residues Trp-102, Glu-103, and Trp-166 contribute side-chain contacts that read the chemical surface of the alkylated base and help discriminate the cap from other nucleotides. Third, the ribose–diphosphate portion of the cap is anchored by direct interactions with basic and aromatic residues. Trp-56 packs against the ribose, while Arg-157 and Lys-162 interact with the alpha and beta phosphate oxygen atoms . Arg-157 itself is stabilized by a salt bridge with Asp-90, helping to correctly position the positively charged side chain for phosphate binding. Finally, water-mediated interactions bridge the protein and the 7-methyl-GDP. Two ordered water molecules link Trp-166 to an α-phosphate oxygen atom, and another water molecule connects Arg-112 to the oxygen atom that forms the ester linkage between the two phosphorus atoms. Together, these direct and water-mediated contacts “lock” the 5’ cap into the binding pocket of eIF4E and create a highly specific recognition site for cap-dependent translation initiation.

IV. Activating Site

As is mentioned above, eIF4E is the rate-limiting factor in translation initiation. Therefore, it is an important target in the regulation of gene expression in eukaryotes. Growth factors, hormones, and mitogens are all involved in pathways that boost eIF4E affinity to the 5' cap via phosphorylation of Ser-209 . This increased binding between elF4E and the 5'cap promotes unwinding of the 5' untranslated region and eventaully the exposure of the start codon. EIf4E is therefore a crucial target for gene expression.

V. References

Joshi B, Cai AL, Keiper BD, Minich WB, Mendez R, Beach CM, Stepinski J, Stolarski R, Darzynkiewicz E, Rhoads RE. Phosphorylation of eukaryotic protein synthesis initiation factor 4E at Ser-209. J Biol Chem. 1995 Jun 16;270(24):14597-603. doi: 10.1074/jbc.270.24.14597. PMID: 7782323.

Marcotrigiano J, Gingras AC, Sonenberg N, Burley SK. Cocrystal structure of the messenger RNA 5' cap-binding protein (eIF4E) bound to 7-methyl-GDP. Cell. 1997 Jun 13;89(6):951-61. doi: 10.1016/s0092-8674(00)80280-9. PMID: 9200613.

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