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Nucleic Acids Research, Vol 25, Issue 24 5119-5124, Copyright © 1997 by Oxford University Press

ARTICLES

Single substitutions of phosphorothioates in the HDV ribozyme G73 define regions necessary for optimal self-cleaving activity

NS Prabhu, G Dinter-Gottlieb and PA Gottlieb
Department of Bioscience and Biotechnology, Drexel University, Philadelphia, PA 19104, USA.

Phosphorothioate (NTPalphaS) analogues were incorporated into the HDV genomic ribozyme by transcription with T7 polymerase. The introduction of a sulfur in place of the pro-Rp oxygen at the phosphate 5'to positions A64, A63, A43, U27, G62, C61, C44, C41, C22and C21appeared to inhibit self-cleavage activity of the G73 genomic ribozyme. Except for position C22, elevated levels of Mg2+rescued the reaction to various extents. When the sites were identified in the RNA sequence, they were clustered in three distinct regions that, in the secondary structure models, are predicted to be primarily single-stranded. Two of these regions have been proposed to form extensive interactions that are thought to involve a homopurine base pair. The third region is thought to be directly associated with assembly of the cleavage site.
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K. Salehi-Ashtiani, A. Luptak, A. Litovchick, and J. W. Szostak
A genomewide search for ribozymes reveals an HDV-like sequence in the human CPEB3 gene.
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