Biophysical Studies of eIF4E Cap-binding Protein: Recognition of mRNA 5′ Cap Structure and Synthetic Fragments of eIF4G and 4E-BP1 Proteins

Niedzwiecka, Anna; Marcotrigiano, Joseph; Stępiński, Janusz; Jankowska-Anyszka, Marzena; Wysłouch‐Cieszyńska, Aleksandra; Dadlez, Michał; Gingras, Anne‐Claude; Mak, Paweł; Darżynkiewicz, Edward; Sonenberg, Nahum; Burley, S.K.; Stolarski, Ryszard

doi:10.1016/s0022-2836(02)00328-5

Cited by 360 publications

(677 citation statements)

References 83 publications

(148 reference statements)

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“…The binding affinity is 10-to 40-fold lower than for eIF4E1 (Table 1). Similarly to eIF4E1, a eightfold reduction in affinity was observed for the dinucleotide m 7 GpppG compared with m 7 GTP (20). Although ITC did not detect eIF4E3 binding to GTP and GDP ( Fig.…”

Section: Resultsmentioning

confidence: 84%

“…The backbone root-mean-square deviation (rmsd) for regular secondary structure elements is 1.86 Å for apo-eIF4E3 versus apo-eIF4E1 (8) and 1.60 Å for m 7 GDP-eIF4E3 versus m 7 GTP-eIF4E1 (25). An extra single turn α helix (denoted α0, residues [16][17][18][19][20] is seen in both eIF4E3 structures at the disordered N terminus (residues 1-30) that is not seen in any other eIF4E structure. We detected nuclear Overhauser effects (NOEs) between this helix and the α1 helix for the apo form, but not the m 7 GDP-bound form.…”

Section: Resultsmentioning

confidence: 99%

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eIF4E3 acts as a tumor suppressor by utilizing an atypical mode of methyl-7-guanosine cap recognition

Osborne

Volpon

Kornblatt

et al. 2013

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

104

143

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Recognition of the methyl-7-guanosine (m 7 G) cap structure on mRNA is an essential feature of mRNA metabolism and thus gene expression. Eukaryotic translation initiation factor 4E (eIF4E) promotes translation, mRNA export, proliferation, and oncogenic transformation dependent on this cap-binding activity. eIF4E-cap recognition is mediated via complementary charge interactions of the positively charged m 7 G cap between the negative π-electron clouds from two aromatic residues. Here, we demonstrate that a variant subfamily, eIF4E3, specifically binds the m 7 G cap in the absence of an aromatic sandwich, using instead a different spatial arrangement of residues to provide the necessary electrostatic and van der Waals contacts. Contacts are much more extensive between eIF4E3-cap than other family members. Structural analyses of other cap-binding proteins indicate this recognition mode is atypical. We demonstrate that eIF4E3 relies on this cap-binding activity to act as a tumor suppressor, competing with the growthpromoting functions of eIF4E. In fact, reduced eIF4E3 in high eIF4E cancers suggests that eIF4E3 underlies a clinically relevant inhibitory mechanism that is lost in some malignancies. Taken together, there is more structural plasticity in cap recognition than previously thought, and this is physiologically relevant.M etabolism of mRNA is a complex and highly regulated process dependent on the association of the methyl-7-guanosine (m 7 G) cap structure on the 5′ end of transcripts with appropriate proteins (1-3). In mammalian cells, the two major cap-binding proteins are the nuclear cap-binding complex (CBC) (4) and eukaryotic translation initiation factor 4E (eIF4E) (3). Specific cap recognition is key for the fate of transcripts and impacts processes such as mRNA processing and bulk mRNA export via the CBC or the nuclear export of specific transcripts as well as bulk translation by eIF4E (5). NMR and crystallographic studies reveal that m 7 G cap is specifically recognized by both the CBC and eIF4E via intercalation of the m 7 G cap moiety with the side chains of two aromatic residues, i.e., an aromatic sandwich (4, 6-9). This is an electrostatically driven process relying on the partial positive charge of the m 7 G cap and the negative π-electron clouds of the aromatic residues. The aromatic residues are completely conserved in these cap-binding proteins. This recognition motif is used almost exclusively by proteins that specifically bind the m 7 G cap including eIF4E, CBC, and vaccinia virus protein VP39 (4, 10).Dysregulation of cap-binding proteins can have striking physiological consequences. For instance through its cap-binding activity and subsequent effects on gene expression, eIF4E plays an important role in proliferation and survival (1, 3). Indeed, eIF4E is overexpressed in about 30% of human cancers and its overexpression is oncogenic in cell culture and animal models (2, 11). Mutation of the cap-binding site of eIF4E impairs its activities in translation, mRNA export, and oncogenic transform...

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Section: Resultsmentioning

confidence: 84%

Section: Resultsmentioning

confidence: 99%

eIF4E3 acts as a tumor suppressor by utilizing an atypical mode of methyl-7-guanosine cap recognition

Osborne

Volpon

Kornblatt

et al. 2013

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

104

143

View full text Add to dashboard Cite

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“…Many of these hydrogen bonding interactions include contributions due to direct electrostatic interactions because the interacting groups are ionised. These insights into structural characteristics of cap-eIF4E complexes were mainly obtained from three-dimensional structures by x-ray crystallography [20,21,22] and multidimensional NMR study [23].…”

Section: Accepted Manuscriptmentioning

confidence: 99%

“…At the current state of crystallographic refinement, there is no well-localised electron density for the side chain of Lys-159 [20,21], therefore structural data for this chain must be obtained by molecular modelling methods. Employing the above variants of eIF4E protein and its ligands, we show that including reasonable evaluations of non polar and entropic contributions to the binding free energy, we obtain better agreement with the experimentally determined free energies of binding when SE model is used rather than the vdW model.…”

Section: Accepted Manuscriptmentioning

confidence: 99%

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Poisson–Boltzmann model analysis of binding mRNA cap analogues to the translation initiation factor eIF4E

Szklarczyk

Zuberek

Antosiewicz

2009

Biophysical Chemistry

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This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. A C C E P T E D M A N U S C R I P T ACCEPTED MANUSCRIPT AbstractThe electrostatic free energy of binding of two analogues of the 5'-mRNA cap, differing in size and electric charge, to the wild type and mutated eukaryotic initiation factor eIF4E was computed using the finite difference solutions to the Poisson-Boltzmann equation. Two definitions of the solute-solvent dielectric boundary were used: van der Waals model, solvent exclusion (SE) model. The computed electrostatic energies were supplemented by estimations of the non polar and entropic contributions. A comparison with experimental data for the investigated systems was done. It appears that the SE model with additional contribution fits experimental findings better than the van der Waals model does.

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Chemical Synthesis of Dinucleotide Cap Analogs

Kore¹,

Shanmugasundaram²

2013

CP Nucleic Acid Chemistry

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This unit describes a reliable, efficient and general method for the synthesis of standard cap analog (mCAP), m(7)G[5']ppp[5']G, and anti-reverse cap analog (ARCA), m(7,3')(O)G[5']ppp[5']G. The synthesis of required intermediate m(7)GDP or m(2)(7,3'O)GDP has been achieved through regioselective methylation of the corresponding diphosphate using dimethyl sulfate under aqueous conditions. Then, the coupling reaction of m(7)GDP or m(2)(7,3'O)GDP with ImGMP using ZnCl(2)/DMF system affords the corresponding cap analog in good yields.

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Biophysical Studies of eIF4E Cap-binding Protein: Recognition of mRNA 5′ Cap Structure and Synthetic Fragments of eIF4G and 4E-BP1 Proteins

Cited by 360 publications

References 83 publications

eIF4E3 acts as a tumor suppressor by utilizing an atypical mode of methyl-7-guanosine cap recognition

eIF4E3 acts as a tumor suppressor by utilizing an atypical mode of methyl-7-guanosine cap recognition

Poisson–Boltzmann model analysis of binding mRNA cap analogues to the translation initiation factor eIF4E

Chemical Synthesis of Dinucleotide Cap Analogs

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