Question

2. The graphs below represent various curves obtained in different in vitro bacterial translation experiments in which a mixture of radioactive amino acids are added to a cell-free translation assay. The peptide product was assayed as counts per minute (cpm) that resulted from incorporation of a mixture of radiolabeled (radioactive) amino acids. The arrows represent the times at which various substances were added to the systems. (6 points) Cpm Cpm Cpm Cpm Time Time Time Time Time IV a) The addition of an antibiotic at the start of the assay results in the curve shown in graph III, while addition at a later time yields the curve shown in graph I. Which step in protein synthesis (translation) is this antibiotic most likely blocking? (2 points). b) Which graph (I-V) most likely depicts the addition of met-URNA (methionine-tRNA) to a system that lacks only this component (2 points). c) Which graphs (I-V) most likely depicts the addition of an antibiotic that binds irreversibly to the A site of the ribosome? For this question, more than one choice may apply. Select all that apply (2 points).

Answer 1

a) In the graph III, the addition of antibiotic at time = 0 leading to no peptide product formation. Similarly in case of graph I, the addition of antibiotic in later stage also inhibit peptide production but slowly or after some time, which suggests that the antibiotic inhibits translation at initiation stage.

b) Graph IV only denotes that met-tRNA was lacking and peptide product is forming only due to addition of it .

c) Graph II and V depicts that antibiotic binds irreversibly to the A site as in both cases the peptide product formation suddenly stops due to addition of antibiotic.