Figure 10.
Both domain III of EF1α and EBS of Bni1p inhibit EF1α/F-actin binding to mRNA in vitro. Bundles containing 100 nM EF1α and 200 nM F-actin were immobilized in the wells of an ELISA plate (lanes 1–3). GST-EBS of Bni1p (100 nM, lane 4) or GST (100 nM, lane 5) was immobilized in the wells as controls. The wells were then blocked with BSA. After blocking, buffer alone (for lanes 1, 4, and 5) or 5 μM recombinant domain III of EF1α (lane 2) or GST-EBS of Bni1p (lane 3) was added and incubated for 1 h. Biotin-labeled β-actin mRNA (3 nM) was then added and processed as usual. Lanes 4 and 5 show that neither GST-EBS of Bni1p nor GST alone binds to the mRNA. Bars, SD (n = 3).