FIGURE 5.
Effect of the sequence of the first three RNA nucleotides on reactivity with HpRppH in vitro. A, position 1. The reactivity of A4AGAA and G4GGAA was compared as in Fig. 4. The subscript in each RNA name indicates the sequence of the four unpaired nucleotides at the 5′ end. Consequently, A4AGAA was equivalent to A4. B and C, position 2. The reactivity of A4AGAA, A4ACAA, and A4AUAA and of G4GGAA, G4GAAA, G4GCAA, and G4GUAA was compared. Although both radioactivity and fluorescence were measured, only the former is shown in the gel images. To avoid modifying the second nucleotide, A4AUAA and G4GUAA were not labeled with fluorescein; instead, the fluorescence of fluorescein-labeled A8XL was used to normalize the data from each time point. The synthesis of A4AAAA was not successful. D, position 3. The reactivity of A4AGAA, A4AGGA, A4AGCA, and A4AGUA was compared. To avoid modifying the third nucleotide, A4AGUA was not labeled with fluorescein. The S.D. of each measurement is reported in supplemental Table S1.