Journal Article

The catalytic efficiency of yeast ribonuclease III depends on substrate specific product release rate

Marc-Andre Comeau, Daniel A. Lafontaine and Sherif Abou Elela

in Nucleic Acids Research

Volume 44, issue 16, pages 7911-7921
Published in print September 2016 | ISSN: 0305-1048
Published online June 2016 | e-ISSN: 1362-4962 | DOI:

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  • Enzymology
  • Molecular Biology and Genetics


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Members of the ribonuclease III (RNase III) family regulate gene expression by triggering the degradation of double stranded RNA (dsRNA). Hundreds of RNase III cleavage targets have been identified and their impact on RNA maturation and stability is now established. However, the mechanism defining substrates’ reactivity remains unclear. In this study, we developed a real-time FRET assay for the detection of dsRNA degradation by yeast RNase III (Rnt1p) and characterized the kinetic bottlenecks controlling the reactivity of different substrates. Surprisingly, the results indicate that Rnt1p cleavage reaction is not only limited by the rate of catalysis but can also depend on base-pairing of product termini. Cleavage products terminating with paired nucleotides, like the degradation signals found in coding mRNA sequence, were less reactive and more prone to inhibition than products having unpaired nucleotides found in non-coding RNA substrates. Mutational analysis of U5 snRNA and Mig2 mRNA confirms the pairing of the cleavage site as a major determinant for the difference between cleavage rates of coding and non-coding RNA. Together the data indicate that the base-pairing of Rnt1p substrates encodes reactivity determinants that permit both constitutive processing of non-coding RNA while limiting the rate of mRNA degradation.

Journal Article.  7172 words.  Illustrated.

Subjects: Enzymology ; Molecular Biology and Genetics