Reduced turnover of the elongation factor EF-1 X ribosome complex after treatment with the protein synthesis inhibitor II from barley seeds.
The effect of the protein synthesis inhibitor II from barley seeds (Hordeum sp.) on protein synthesis was studied in rabbit reticulocyte lysates. Inhibitor treatment of the lysates resulted in a rapid decrease in amino acid incorporation and an accumulation of heavy polysomes, indicating an effect of the inhibitor on polypeptide chain elongation. The protein synthesis inhibition was due to a catalytic inactivation of the large ribosomal subunit with no effect on the small subparticle. The inhibitor-treated ribosomes were fully active in participating in the EF-1-dependent binding of [14C]phenylalanyl-tRNA to poly(U)-programmed ribosomes in the presence of GTP and the binding of radioactively labelled EF-2 in the presence of GuoPP[CH2]P. Furthermore, the ribosomes were still able to catalyse peptide-bond formation. However, the EF-1- and ribosome-dependent hydrolysis of GTP was reduced by more than 40% in the presence of inhibitor-treated ribosomes, while the EF-2- and ribosome-dependent GTPase remained unaffected. This suggests that the active domains involved in the two different GTPases are non-identical. Treatment of reticulocyte lysates with the barley inhibitor resulted in a marked shift of the steady-state distribution of the ribosomal phases during the elongation cycle as determined by the ribosomal content of elongation factors. Thus, the content of EF-1 increased from 0.38 mol/mol ribosome to 0.71 mol/mol ribosome, whereas the EF-2 content dropped from 0.20 mol/mol ribosome at steady state to 0.09 mol/mol ribosome after inhibitor treatment. The data suggest that the inhibitor reduces the turnover of ribosome-bound ternary EF-1 X GTP X aminoacyl-tRNA complexes during proof-reading and binding of the cognate aminoacyl-tRNA by inhibiting the EF-1-dependent GTPase.
D000818:Animals; D020963:GTP Phosphohydrolase-Linked Elongation Factors / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000378:metabolism; D001467:Hordeum; D046911:Macromolecular Substances; D010441:Peptide Chain Elongation, Translational / Q000187:drug effects; D020648:Peptide Elongation Factor 1; D020652:Peptide Elongation Factor 2; D010445:Peptide Elongation Factors / Q000378:metabolism*; D010458:Peptidyl Transferases / Q000378:metabolism; D010936:Plant Extracts / Q000494:pharmacology*; D037121:Plant Lectins; D011817:Rabbits; D051381:Rats; D012156:Reticulocytes / Q000378:metabolism; D012270:Ribosomes / Q000187:drug effects* / Q000378:metabolism; D012276:Ricin / Q000494:pharmacology; D012639:Seeds
D046911:Macromolecular Substances; D020648:Peptide Elongation Factor 1; D020652:Peptide Elongation Factor 2; D010445:Peptide Elongation Factors; D010936:Plant Extracts; D037121:Plant Lectins; D006160:Guanosine Triphosphate; D012276:Ricin; D010458:Peptidyl Transferases; D020963:GTP Phosphohydrolase-Linked Elongation Factors