Figure 3.

Effect of antibiotics on in vitro incorporation of amino acids by stalled complex of ribosomes from S. griseus and S. collinus.
Cells from the exponential phase of growth were incubated in absence (▪) or presence (□) of sub‐inhibitory concentrations of produced antibiotics. In S. griseus with 3.5 µM streptomycin (A) and S. collinus with 5 µM kirromycin (B). At indicated time intervals samples were taken for preparation of S30 fractions. Pre‐incubated S30 fractions (2 A₂₆₀ units) were used for translation of poly(U) in reaction mixtures (100 µl) containing 50 mM Tris‐HCl pH 7.6, 10 mM MgCl₂, 80 mM NH₄Cl, 1 mM dithiothreitol, 5 mM phosphoenol pyruvate, 1 mM ATP, 0.5 mM GTP, 15 mM phenylalanine and 100 µg poly(U). After 15 min of incubation, phenylalanine incorporation was saturated. To the stalled complex of ribosomes was added 20 µM each of U¹⁴C‐l‐alanine, U¹⁴C‐l‐arginine, U¹⁴C‐l‐leucine, U¹⁴C‐l‐lysine and 50 µM each of remaining unlabelled amino acids. These mixtures were incubated at 32°C for 15 min. Reactions were terminated by trichloroacetic acid, boiled at 95°C for 20 min and filtered over Whatmam GF/C filters and assayed for radioactivity. The sensitivity of ribosomes of the 10 h cultures to the presence of produced antibiotics was examined at the translation of poly(U). Dialysed and pre‐incubated S30 fractions of S. griseus (C) and S. collinus (D) were incubated in the reaction mixtures with U¹⁴C‐l‐phenylalanine (0.2 µCi) and antibiotics as indicated. Radioactivity in the hot trichloroacetic acid insoluble fractions was measured by liquid scintillation counter. The values are average from four independent experiments.