Correlation between the distribution of the reversing factor and eukaryotic initiation factor 2 in heme‐deficient or double‐stranded RNA‐inhibited reticulocyte lysates

FEBS Letters

Thomas, N.Shaun B.¹  Hurst, Robin²  London, Irving M.¹  Matts, Robert L.² 

[1] Harvard-Massachusetts Institute of Technology, Division of Health Sciences and Technology, Massachusetts Institute of Technology, Cambridge, MA 02139, USA;Department of Biochemistry, Oklahoma State University, Stillwater, OK 74078, USA

关键词: Protein synthesis;    Reticulocyte lysate;    Reversing factor;    Eukaryotic initiation factor 2;    60 S ribosome;    Ribosomal subunit binding;    eIF-2;    eukaryotic initiation factor 2;    eIF-2α;    α(38 kDa)-subunit of eIF-2;    eIF-2(α-P);    eIF-2 phosphorylated on α-subunit;    eIF-2 · GDP;    binary complex of eIF-2 with GDP;    eIF-2(α-P) · GDP;    binary complex of eIF-2 with GDP;    phosphorylated on the α-subunit of eIF-2;    RF;    reversing factor (also referred to in the literature as GEF and eIF-2B);    RF · eIF-2(α-P);    complex of RF with eIF-2 phosphorylated on its α-subunit;    HRI;    heme-regulated eIF-2α kinase;    dsRNA;    double-stranded RNA;    dsI;    dsRNA-activated eIF-2α kinase;    SDS-PAGE;    SDS-polyacrylamide gel electrophoresis;   

DOI  :  10.1016/0014-5793(88)80310-7

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

The recycling of eukaryotic initiation factor eIF-2 requires the exchange of GDP for GTP, in a reaction catalyzed by the reversing factor (RF). Recent studies have suggested that a 60 S ribosomal subunit-bound eIF-2 · GDP complex is an intermediate in protein chain initiation. We have monitored the distribution of RF in heme-deficient and dsRNA-inhibited lysates by immunoblot analysis of sucrose gradient fractions and have compared the distribution with that of eIF-2(α-³²P). RF and eIF-2(αP) were both found to be tightly associated with 60 S and 80 S ribosomes, as their distribution did not change in gradients containing up to 0.1 M K⁺. The association of eIF-2(α-³²P) and RF with 60 S and 80 S ribosomes was enhanced in the presence of F⁻, indicating the presence of an endogenous ribosome-associated phosphatase activity which is capable of dephosphorylating eIF-2(αP) in the absence of F⁻. These observations are consistent with the hypothesis that under physiologic conditions, RF interacts with the 60 S-bound eIF-2 · GDP complex to promote the dissociation of GDP from eIF-2 and the release of eIF-2 from the 60 S subunit as a complex with RF.

【 授权许可】

Unknown