The addition of Pb2+ to a small RNA molecule consisting of an asymmetric internal loop of six nucleotides results in site-specific cleavage followed by hydrolysis of the 2′,3′-cyclic phosphate intermediate [Pan, T., & Uhlenbeck, O. C. (1992) Nature 358, 560–563]. Here we show that the reaction is highly specific for Pb2+ and the cleavage rate increases exponentially with pH from 5.5 to 7.0, both in the presence and in the absence of Mg2+. This suggests that the reaction mechanism involves Pb2+ hydroxide acting as a base. Several sequence variants of the RNA are found to be equally active in both steps of the reaction, suggesting that they fold into a similar structure.
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