NH-aIF1 stimulates binding of RNAo to S. solfataricus 30S subunits. (A) S. solfataricus ribosomes (0.1 μM) were incubated with a/eIF2·GTP·Met-tRNAi (lanes 2,4–7), radioactively labeled RNAo (0.01 μM) (lanes 1–7) and increasing amounts of aIF1 (0.1, 0.2, 0.5 and 1 μM, respectively) (lanes 4–7) at 70°C. The samples were loaded on a native 4% polyacrylamide gel and the radioactive bands were visualized using a PhosphorImager. (Lane 1) Only RNAo (0.01 μM) was incubated with ribosomes (0.1 μM). (Lanes 2,3) Ribosomes were incubated with RNAo in the presence of a/eIF2·GTP·Met-tRNAi or aIF1 (1μM), respectively. The position of the ribosomal subunits was determined by Coomassie staining of the gel as described in the legend to Figure 4. (B) A graphical representation of two independent experiments of RNAo binding to ribosomes as a function of the aIF1 concentration is shown. The error bars represent standard deviations.
