Cap binding protein complex that restores protein synthesis in heat-shocked Ehrlich cell lysates contains highly phosphorylated eIF-4E.
Cell-free protein-synthesizing systems derived from Ehrlich ascites tumor cells that have been exposed to elevated temperatures retain the inhibition of translation that is seen at the cellular level. A multisubunit cap binding protein complex able to restore protein synthesis in these cell free systems was purified from Ehrlich ascites tumor cells via affinity chromatography using m7GTP-Sepharose and fast protein liquid chromatography on Mono Q. The purified complex contains an Mr 220,000 polypeptide (p220) and an Mr 28,000 polypeptide (p28), both of which are components of eukaryotic initiation factor 4F (eIF-4F). p28 is identical to eIF-4E. Restoring activity was relatively free of the Mr 46,000 polypeptide (p46) that is the third component of eIF-4F and does not appear to be dependent on its presence. p28 associated in a complex with p220 is 85% phosphorylated; however, the majority of p28 is not associated with p220, and this free form is only about 50% phosphorylated. The correlation between association of p28 with p220 and high levels of p28 phosphorylation suggests a possible role for phosphorylation in association of p220 with p28.
Lamphear
B J
BJ
Department of Biochemistry and Cancer Center, University of Rochester, New York 14642.
Panniers
R
R
D000818:Animals; D002286:Carcinoma, Ehrlich Tumor / Q000378:metabolism*; D002352:Carrier Proteins / Q000302:isolation & purification / Q000378:metabolism*; D002846:Chromatography, Affinity; D002851:Chromatography, High Pressure Liquid; D039561:Eukaryotic Initiation Factor-4E; D039562:Eukaryotic Initiation Factor-4F; D006358:Hot Temperature; D007525:Isoelectric Focusing; D008970:Molecular Weight; D010448:Peptide Initiation Factors / Q000302:isolation & purification / Q000378:metabolism*; D010766:Phosphorylation; D014176:Protein Biosynthesis; D039381:RNA Cap-Binding Proteins
D002352:Carrier Proteins; D039561:Eukaryotic Initiation Factor-4E; D039562:Eukaryotic Initiation Factor-4F; D010448:Peptide Initiation Factors; D039381:RNA Cap-Binding Proteins