Internal and overall motions of the translation factor eIF4E: cap binding and insertion in a CHAPS detergent micelle.
The mRNA cap-binding protein eIF4E is the limiting factor in the eIF4F translation initiation complex, which mediates the binding of the 40S ribosome to the mRNA. 15N relaxation studies have been used to characterize the backbone dynamics of deuterated eIF4E in a CHAPS micelle for the apoprotein, the m7GDP-bound form, and the dinucleotide (m7GpppA)-bound form, as well as for CHAPS-free eIF4E. Large differences in overall correlation time between the CHAPS-free form (11.8 ns) and samples containing different concentrations of CHAPS (15.9-19.4 ns) indicate that eIF4E is embedded in a large micelle in the presence of CHAPS, with a total molecular weight in the range of 40-60 kDa. CHAPS seems to restrict the mobility of the a2-b3 and a4-b5 loops which are thought to be embedded in the micelle. No significant changes in overall mobility were seen between the m7 GDP-bound form, the m7GpppA-bound form, and the apoprotein. Amide hydrogen exchange data indicate the presence of slowly exchanging amides in two surface-exposed helices (a2 and a4), as well as the a4-b5 loop, indicating protection by the CHAPS micelle. The micelle covers the convex side of the protein away from the cap-binding site.
McGuire
A M
AM
Graduate Program in Biophysics, Harvard Medical School, Boston, MA 02115, USA.
Matsuo
H
H
Wagner
G
G
D002793:Cholic Acids; D003001:Cloning, Molecular; D003902:Detergents; D039561:Eukaryotic Initiation Factor-4E; D006153:Guanosine Diphosphate / Q000031:analogs & derivatives; D006860:Hydrogen Bonding; D007700:Kinetics; D008823:Micelles; D008958:Models, Molecular; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods; D009690:Nucleic Acid Conformation; D010448:Peptide Initiation Factors / Q000737:chemistry* / Q000378:metabolism; D017433:Protein Structure, Secondary; D012314:RNA Cap Analogs / Q000737:chemistry / Q000378:metabolism; D012315:RNA Caps / Q000737:chemistry* / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000378:metabolism
D002793:Cholic Acids; D003902:Detergents; D039561:Eukaryotic Initiation Factor-4E; D008823:Micelles; D010448:Peptide Initiation Factors; D012314:RNA Cap Analogs; D012315:RNA Caps; D011994:Recombinant Proteins; C111106:7-methylguanosine 5'-diphosphate; D006153:Guanosine Diphosphate; C028213:3-((3-cholamidopropyl)dimethylammonium)-1-propanesulfonate