Figure 5.

(A) Scheme for interference rescue that requires a second mutation for maximum activity. Unlike the scheme in Fig. 5, activity of the G to A point mutant is not significantly rescued by DMPαS substitution unless a second nucleotide is also changed. In this case, U −1 must be mutated to a C. This restores the wobble configuration of the base pair. (B) A portion of the sequencing autoradiogram showing the reaction of L+1 ScaI DMPαS or AMPαS at 50°C in 4 mM MgCl₂. Reactions with dT(-1)P and dC(-1)P were for 2 and 10 min, respectively. The magnitude of the interference rescue was estimated by comparing the intensity of each band in the DMPαS lane to the intensity in the AMPαS lane and normalizing to the intensity at all other positions in the RNA.