Interaction of three Caenorhabditis elegans isoforms of translation initiation factor eIF4E with mono- and trimethylated mRNA 5' cap analogues.
Translation initiation factor eIF4E binds the m(7)G cap of eukaryotic mRNAs and mediates recruitment of mRNA to the ribosome during cap-dependent translation initiation. This event is the rate-limiting step of translation and a major target for translational control. In the nematode Caenorhabditis elegans, about 70% of genes express mRNAs with an unusual cap structure containing m(3)(2,2,7)G, which is poorly recognized by mammalian eIF4E. C. elegans expresses five isoforms of eIF4E (IFE-1, IFE-2, etc.). Three of these (IFE-3, IFE-4 and IFE-5) were investigated by means of spectroscopy and structural modelling based on mouse eIF4E bound to m(7)GDP. Intrinsic fluorescence quenching of Trp residues in the IFEs by iodide ions indicated structural differences between the apo and m(7)G cap bound proteins. Fluorescence quenching by selected cap analogues showed that only IFE-5 forms specific complexes with both m(7)G- and m(3)(2,2,7)G-containing caps (K(as) 2 x 10(6) M(-1) to 7 x 10(6) M(-1)) whereas IFE-3 and IFE-4 discriminated strongly in favor of m(7)G-containing caps. These spectroscopic results quantitatively confirm earlier qualitative data derived from affinity chromatography. The dependence of K(as) on pH indicated optimal cap binding of IFE-3, IFE-4 and IFE-5 at pH 7.2, lower by 0.4 pH units than that of eIF4E from human erythrocytes. These results provide insight into the molecular mechanism of recognition of structurally different caps by the highly homologous IFEs.
Stachelska
Alicja
A
Department of Physics and Biophysics, University of Warmia and Mazury in Olsztyn, Olsztyn, Poland.
Wieczorek
Zbigniew
Z
Ruszczyńska
Katarzyna
K
Stolarski
Ryszard
R
Pietrzak
Monika
M
Lamphear
Barry J
BJ
Rhoads
Robert E
RE
Darzynkiewicz
Edward
E
Jankowska-Anyszka
Marzena
M
D000818:Animals; D017173:Caenorhabditis elegans / Q000378:metabolism*; D029742:Caenorhabditis elegans Proteins / Q000737:chemistry / Q000378:metabolism*; D039561:Eukaryotic Initiation Factor-4E / Q000737:chemistry / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007454:Iodides / Q000737:chemistry; D007700:Kinetics; D008745:Methylation; D008958:Models, Molecular; D020033:Protein Isoforms / Q000378:metabolism; D017434:Protein Structure, Tertiary; D012315:RNA Caps / Q000737:chemistry / Q000378:metabolism*; D012333:RNA, Messenger / Q000737:chemistry / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013050:Spectrometry, Fluorescence / Q000379:methods; D013056:Spectrophotometry, Ultraviolet; D040681:Structural Homology, Protein; D014364:Tryptophan / Q000737:chemistry
D029742:Caenorhabditis elegans Proteins; D039561:Eukaryotic Initiation Factor-4E; D007454:Iodides; D020033:Protein Isoforms; D012315:RNA Caps; D012333:RNA, Messenger; D011994:Recombinant Proteins; D014364:Tryptophan