Pum2 W344 mediates cap binding and translational repression. (A) Pum2 deletion mutants and a W344G point mutant were expressed in reticulocyte lysates and applied to m7G-Sepharose; the percent bound was determined by Western blotting. (B) Sequence alignment of a selected region of Pum2 protein among animal species. The underlined W (corresponding to W344 in Xenopus laevis) is conserved among these and probably most other vertebrates as well. (C) Pum2 WT, Pum2 W344G, eIF4E, and ePAB were expressed in reticulocyte lysates, applied to GDP-Sepharose or m7G-Sepharose, and proteins that were retained were analyzed by Western blotting. (D) Recombinant Pum2 WT and W344G were applied to m7G-Sepharose and the bound material analyzed by Western blotting.
