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Reconstitution of a Group I Intron Self-Splicing Reaction with an Activator RNA

Gerda van der Horst, Andreas Christian and Tan Inoue
Proceedings of the National Academy of Sciences of the United States of America
Vol. 88, No. 1 (Jan. 1, 1991), pp. 184-188
Stable URL: http://www.jstor.org/stable/2355734
Page Count: 5
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Since scans are not currently available to screen readers, please contact JSTOR User Support for access. We'll provide a PDF copy for your screen reader.
Reconstitution of a Group I Intron Self-Splicing Reaction with an Activator RNA
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Abstract

The self-splicing rRNA intron of Tetrahymena thermophila belongs to a subgroup of group I introns that contain a conserved extra stem-loop structure termed P5abc. A Tetrahymena mutant precursor RNA lacking this P5abc is splicing-defective under standard conditions (5 mM MgCl2/200 mM NH4Cl, pH 7.5) in vitro. However, the mutant precursor RNA by itself is capable of performing the self-splicing reaction without P5abc under different conditions (15mM MgCl2/2 mM spermidine, pH 7.5). We have investigated the functional role of the P5abc in the mechanism of the self-splicing reaction. When an RNA consisting of the P5abc but lacking the rest of the Tetrahymena intron is incubated with the mutant precursor, the self-splicing reaction proceeds highly efficiently under standard conditions (5 mM MgCl2/200 mM NH4Cl, pH 7.5). Two steps of the bimolecular self-splicing reaction can be performed accurately by a shortened precursor RNA containing all essential components required in the self-splicing reaction and an activator RNA consisting of the P5abc. Gel-mobility-shift assays suggest that two molecules associate by a direct RNA·RNA interaction during the splicing reaction. The results imply that there might exist other small RNAs whose role is to activate ribozymes.

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