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Nucleic Acids Research 2006 34(2):584-592; doi:10.1093/nar/gkj463
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Published online 23 January 2006

© The Author 2006. Published by Oxford University Press. All rights reserved
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Article

Unbiased in vitro selection reveals the unique character of the self-cleaving antigenomic HDV RNA sequence

Atef Nehdi and Jean-Pierre Perreault*

RNA Group/Groupe ARN, Département de Biochimie, Faculté de médecine et des sciences de la santé, Université de Sherbrooke Sherbrooke, Québec, Canada J1H 5N4

*To whom correspondence should be addressed. Tel: +1 819 564 5310; Fax: +1 819 564 5340; Email: Jean-Pierre.Perreault{at}USherbrooke.ca

Received December 2, 2005. Revised January 6, 2006. Accepted January 6, 2006.

In order to revisit the architecture of the catalytic center of the antigenomic hepatitis delta virus (HDV) ribozyme we developed an unbiased in vitro selection procedure that efficiently selected novel variants from a relatively small set of sequences. Using this procedure we examined all possible variants from a pool of HDV ribozymes that had been randomized at 25 positions (425). The isolated set of sequences shows more variability than do the natural variants. Nucleotide variations were found at all randomized positions, even at positions when the general belief was that the specific base was absolutely required for catalytic activity. Covariation analysis supports the presence of several base pairs, although it failed to propose any new tertiary contacts. HDV ribozyme appears to possess a greater number of constraints, in terms of sequences capable of supporting the catalysed cleavage, than do other catalytic RNAs. This supports the idea that the appearance of this catalytic RNA structure has a low probability (i.e. is a rare event), which may explain why to date it has been found in nature only in the HDV. These contrasts with the hammerhead self-cleaving motif that is proposed to have multiple origins, and that is widespread among different organisms. Thus, just because a self-cleaving RNA motif is small does not imply that it occurs easily.


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