| Abstrakt: |
Ribosomal proteins S 12, S5 and S4 ofEscherichia coli are essential for the control of translational accuracy. Their yeast equivalents, i.e., S28, S4 and S13, have also been implicated in this process. Using a poly(U)-dependent cell-free translation system, we determined the accuracy of translation and the sensitivity to antibiotic paromomycin of yeast ribosomes carrying mutant ribosomal proteins S28 and/or S4. Our results confirm by quantitative biochemical methods previous genetic data showing that proteins S28 and S4 are involved in the decoding activity of the ribosome and interact to control translational accuracy. We find that the suppressor mutationSUP44 in yeast S4, decreased the accuracy of translation. To examine the effect of mutant S28, we disruptedRPS28B and introduced inRPS28A the same substitutions that cause hyperaccurate translation or antibiotic resistance in bacteria. Three of these substitutions (Lys-62 → Asn, Thr or Gin) similarly increased translational accuracy in vitro or antibiotic resistance. In the presence of theSUP44 mutation, these substitutions partially reversed the decrease of translational accuracy caused bySUP44. However, the Lys-62 →Arg substitution decreased translational accuracy and caused antibiotic sensitivity both in nonsuppressor and inSUP44 haploids. These results establish the role of Lys-62 of S28 in optimizing translational accuracy and provide a more precise view of the functional role of two important ribosomal proteins. |
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