FIG. 5.
LaDN inhibits the initiation of formation of 80S ribosome complexes. RNA-ribosome binding assay was performed in a cycloheximide (0.6 mM)-treated HeLa cell extract (40 μl) with poliovirus (∼105 cpm) (A), HCV IRES-poliovirus chimera (∼106 cpm) (B), and rabbit globin (∼105 cpm) (C) 3′-end-labeled mRNAs. The extract was incubated with GST-La226-348 (4 μg) or GST (4 μg) at 34°C for 15 min and diluted with ice-cold high-salt buffer (66). Ribosome complexes were analyzed as described in Materials and Methods.