Question

A reticulate lysate is a system that contains all the components required for translation (ribosomal subunits, tRNA, amino acids, etc) and allows the reaction to be carried out in a test tube. The polypeptide that is synthesized from the system depends on the sequence of the RNA. For example adding the RNA 5’AUGUUUUUUUUUUUU to the lysate directs the synthesis of Met-Phe-Phe-Phe-Phe.

10a. If you add an antibiotic stops translation by specifically blocking the translocation of tRNA from the A site to the P site of the ribosome to the system, what will be the sequence of polypeptide produced by the same RNA? Explain your answer briefly.

10b. You add a new antibiotic with unknown function to the lysate and no polypeptide was produced. Suggest a step in translation that is inhibited by the antibiotics.

Answer 1

a. Met-Phe. This is because the first codon AUG codes for Met and the first t-RNA sits at the P site, the next t-RNA will sit at A site site and add another amino acid (Phe as per the sequence of RNA) to the polypeptide chain. After this the first t-RNA leaves P site and the second t-RNA should move to A site to facilitate the other t-RNA to come but since A to P movement is inhibited, translation stops here.

b. Since no polypeptide was produced, there could be multiple reasons. The antibiotic inhibited start codon translation, maybe the t-RNA attachment was inhibited. Mostly, the antibiotic must be inhibiting the ribosome from attaching to the mRNA fragment.

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