FIGURE 3.

hDcp2 does not utilize the cap structure as substrate. (A) hDcp2 is unable to hydrolyze the cap structure in the absence of the RNA body. ³²P-labeled cap analog was incubated with 10 pmole of each hDcp2 protein and the products resolved by TLC. The scavenger decapping activity DcpS, which can hydrolyze the cap structure, was used as the positive control (lane 2). Standards are indicated at left. (B) hDcp2 is unable to cross-link to cap analog. ³²P-labeled cap analog was UV cross-linked with 20 pmole of the indicated proteins and the products resolved by 12.5% SDS-PAGE. GST fusion protein of the cap-binding protein, eIF4E (lane 1), was used as the positive control. Histidine-tagged hDcp2 protein in lane 2, and an unrelated RNA-binding protein control, GST-mDAZL (Jiao et al. 2002), was used in lane 3. Molecular weight standards are indicated at right. The cap analog substrate is shown at bottom, and the star denotes the labeled phosphate.