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Nucleic Acids Research, 1993, Vol. 21, No. 20 4685-4689
© 1993

MOLECULAR BIOLOGY

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

Yo Kikuch, Noriko Sasaki-Tozawa and Kyoko Suzuki

Mitsubishi Kasei Institute of Life Sciences 11 Minamiooya, Machida-shi, Tokyo 194, Japan

Received July 22, 1993. Revised August 27, 1993. Accepted August 27, 1993.

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 RNApre- 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-pretRNA 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 pretRNA- 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.


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