Studies on the role of eukaryotic nucleotide exchange factor in polypeptide chain initiation

D. J. Goss, Lawrence J Parkhurst, H. B. Mehta, C. L. Woodley, A. J. Wahba

Research output: Contribution to journalArticle

36 Citations (Scopus)

Abstract

Interactions of eukaryotic 5-dimethylaminonaphthalene-1-sulfonyl-initiation factor 2 (eIF-2) from rabbit reticulocytes and the guanine nucleotide exchange factor (GEF), Met-tRNA(f), GTP, and GDP were monitored by changes in fluorescence anisotropy and radioactive filtration assays. At 1 mM Mg2+, radioactive filtration assays demonstrate that GEF is necessary for nucleotide exchange. We did not observe a GDP dependence in the association reaction of eIF-2.GEF for GDP concentrations from 0.01 to 20 μM. This is in disagreement with the model: eIF-2.GDP + GEF of GTP caused a decrease in fluorescence anisotropy which is interpreted as a dissociation of eIF-2.GEF. We propose an asymmetrical model of ternary complex (eIF-2.GTP.Met-tRNA(f)) formation where 1) GDP does not displace GEF and 2) GTP replaces GEF and presumably GDP. For reticulocyte eIF-2, phosphorylation of the α subunit greatly inhibits protein synthesis. This inhibition derives neither from failure of GEF to bind to eIF-2(αP) nor from greatly enhanced binding of GEF. The inhibition results from the requirement of very high levels of GTP (100 μM) to dissociate the eIF-2(αP).GEF complex.

Original languageEnglish (US)
Pages (from-to)7374-7377
Number of pages4
JournalJournal of Biological Chemistry
Volume259
Issue number12
StatePublished - Aug 23 1984

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Guanine Nucleotide Exchange Factors
Prokaryotic Initiation Factor-2
Nucleotides
Peptides
Guanosine Triphosphate
Fluorescence Polarization
Reticulocytes
Assays
Anisotropy
Fluorescence
Phosphorylation
Association reactions
Rabbits

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Studies on the role of eukaryotic nucleotide exchange factor in polypeptide chain initiation. / Goss, D. J.; Parkhurst, Lawrence J; Mehta, H. B.; Woodley, C. L.; Wahba, A. J.

In: Journal of Biological Chemistry, Vol. 259, No. 12, 23.08.1984, p. 7374-7377.

Research output: Contribution to journalArticle

Goss, D. J. ; Parkhurst, Lawrence J ; Mehta, H. B. ; Woodley, C. L. ; Wahba, A. J. / Studies on the role of eukaryotic nucleotide exchange factor in polypeptide chain initiation. In: Journal of Biological Chemistry. 1984 ; Vol. 259, No. 12. pp. 7374-7377.
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