Figure 2.

Cleavage by wild-type M1 RNA in the presence of different divalent metal ion combinations. (a) Diagram showing miscleavage of pATSerCG at the −1 position by wild-type M1 RNA in the presence of different combinations of divalent metal ions as indicated. No miscleavage at −1 was detected when cleavage was performed in the presence of the given Mg²⁺/Sr²⁺ combinations. The given values are averages of several independent experiments, and experimental errors are indicated in the figure. (b) Cleavage of [α-³²P]UTP internally labeled pATSerCG (except the last three lanes to the right in which we used [γ-³²P]ATP 5′ end-labeled pATSerUA, explaining why the 5′-matured cleavage product is not seen) as a function of different divalent metal ion combinations as indicated. In the experiments where two Me²⁺ were mixed the final concentration of each Me²⁺ was 20 mM, resulting in a total final divalent metal ion concentration of 40 mM. Time represents the time of incubation of M1 RNA in the presence of pATSerCG (or pATSerUA). The controls were: Ctrl H₂O, incubation of pATSerCG in the presence of only H₂O for 180 min; Ctrl Zn/Co, incubation of pATSerUA in the presence of 20 mM Zn²⁺ 20 mM Co(NH₃)₆³⁺, and no M1 RNA; Co, Co(NH₃)₆³⁺. Note that pATSerUA was cleaved only at the +1 position, whereas pATSerCG was cleaved both at the correct position +1 and at −1 (see also ref. 15).