Kinetic determination of the effects of ADP-ribosylation on the interaction of eukaryotic elongation factor 2 with ribosomes

J Biol Chem. 1990 Apr 15;265(11):6030-4.


The effect of ADP-ribosylation on the function of eukaryotic elongation factor 2 (EF-2) was investigated by kinetic analysis of the EF-2-catalyzed hydrolysis of GTP in the presence of ribosomes and by direct determination of the affinity of the modified factor for the ribosome. Under conditions where the concentration of EF-2 was rate-limiting, the ADP-ribosylation reduced the maximum rate of GTP hydrolysis and the second order rate constant Kcat/Km by approximately 50%. A similar decrease in Kcat and Kcat/Km was observed when the concentration of ribosomes were kept rate-limiting. The affinity of EF-2 for the pretranslocation type of ribosomes was reduced by 2 orders of magnitude after ADP-ribosylation. No effect was observed in the interaction with the post-translocation type of ribosomes, the ribosomal conformation responsible for activation of the EF-2-dependent GTPase. We conclude that the ADP-ribosylation affects both the association of the modified factor with pretranslocation ribosomes and the hydrolytic capacity of the factor.

MeSH terms

  • Adenosine Diphosphate Ribose / metabolism*
  • Animals
  • Carcinoma, Ehrlich Tumor / metabolism
  • Diphtheria Toxin / pharmacology
  • Guanosine Triphosphate / metabolism
  • Kinetics
  • Mice
  • NAD / metabolism*
  • Peptide Elongation Factor 2
  • Peptide Elongation Factors / metabolism*
  • Phosphoproteins / metabolism
  • Ribosomes / drug effects
  • Ribosomes / metabolism*
  • Ricin / pharmacology


  • Diphtheria Toxin
  • Peptide Elongation Factor 2
  • Peptide Elongation Factors
  • Phosphoproteins
  • NAD
  • Adenosine Diphosphate Ribose
  • Guanosine Triphosphate
  • Ricin