Figure 6.

Binding of ³⁵S-labeled recombinant eIF6 to 60S ribosomal subunits. Reaction mixtures (100 μl each) contained 20 mM Tris⋅HCl, pH 7.5, 100 mM KCl, 1 mM MgCl₂, 1 mM DTT, and 2.0 A₂₆₀ unit of either 40S or 60S ribosomal subunits. Reactions were initiated by the addition of 320 ng (≈13 pmol) of purified ³⁵S-labeled recombinant eIF6 containing 5,000 cpm of radioactivity. After incubation at 37°C for 5 min, the Mg²⁺ concentration of each reaction was raised to 5 mM, and the reactions were analyzed by sucrose gradient centrifugation as described in the legend to Fig. 5, except that each fraction was assayed for ³⁵S radioactivity by counting in Aquasol in a liquid scintillation spectrometer. •, 60S ribosomal subunits/³⁵S-eIF6; ○, 40S ribosomal subunits/³⁵S-eIF6; ▪, ³⁵S-eIF6 alone.