FIG. 2.

Purified eIF4E inhibits decapping. (A) Analysis of decapping activity by DCP1 alone (left panel) and increasing amounts of GST-eIF4E (purified with a cap affinity column): either 8.3 pmol of GST-eIF4Ep (middle panel [2.5×]) or 16.7 pmol of GST-eIF4Ep (right panel [5×]). Decapping by His-DCP1p was assayed over a 30-min time course with aliquots removed at the designated time points. The products of the reaction were resolved by thin-layer chromatography. (B) Decapping is inhibited upon addition of eIF4E. The histogram shows the amount of decapping activity obtained with increasing amounts of eIF4E. Relative activity was determined by assaying decapping ± eIF4E in multiple experiments and is expressed as the amount of decapping relative to that with DCP1 alone (set at 100%). The 30-min time point is shown on the histogram.