Artificial self-cleaving molecules consisting of a tRNA precursor and the catalytic RNA of RNase P

We synthesized two types of chimeric RNAs between the catalytic RNA subunit of RNase P from Escherichia coli (M1 RNA) and a tRNA precursor (pre-tRNA); one had pre-tRNA at the 3′ side to the M1 RNA (M1 RNA-pre-tRNA). The second had pre-tRNA at the 5′ side of the M1 RNA (pre-tRNA-M1 RNA). Both molecules were self-cleaving RNAs. The self-cleavage of M1 RNA-pre-tRNA occurred at the normal site (5′-end of mature tRNA sequence) and proceeded under the condition of 10 mM Mg²⁺ concentration. This reaction at 10 mM Mg²⁺ was an intramolecular reaction (cis-cleavage), while, at 40 mM and 80 mM Mg²⁺, trans-cleavage partially occurred. The self-cleavage rate was strictly affected by the distance between the M1 RNA and the pre-tRNA in the molecule. The self-cleavage of pre-tRNA-M1 RNA occurred mainly at three sites within the mature tRNA sequence. This cleavage did not occur at 10 mM Mg²⁺. Use of M1 RNA-pre-tRNA molecule for the in vitro evolution of M1 RNA is discussed.