Mechanistic characterization of the HDV genomic ribozyme: The cleavage site base pair plays a structural role in facilitating catalysis

Andrea L. Cerrone-Szakal, Durga M. Chadalavada, Barbara L. Golden, Philip C. Bevilacqua

Research output: Contribution to journalArticlepeer-review

16 Scopus citations


The hepatitis delta virus (HDV) ribozyme occurs in the genomic and antigenomic strands of the HDV RNA and within mammalian transcriptomes. Previous kinetic studies suggested that a wobble pair (G•U or A+•C) is preferred at the cleavage site; however, the reasons for this are unclear. We conducted sequence comparisons, which indicated that while G•U is the most prevalent combination at the cleavage site, G-C occurs to a significant extent in genomic HDV isolates, and G•U, G-C, and A-U pairs are present in mammalian ribozymes. We analyzed the folding of genomic HDV ribozymes by free energy minimization and found that variants with purine-pyrimidine combinations at the cleavage site are predicted to form native structures while pyrimidine-purine combinations misfold, consistent with earlier kinetic data and sequence comparisons. To test whether the cleavage site base pair contributes to catalysis, we characterized the pH and Mg2+-dependence of reaction kinetics of fast-folding genomic HDV ribozymes with cleavage site base pair purine-pyrimidine combinations: G•U, A-U, G-C, and A+•C. Rates for these native-folding ribozymes displayed highly similar pH and Mg 2+ concentration dependencies, with the exception of the A +•C ribozyme, which deviated at high pH. None of the four ribozymes underwent miscleavage. These observations support the A +•C ribozyme as being more active with a wobble pair at the cleavage site than with no base pair at all. Overall, the data support a model in which the cleavage site base pair provides a structural role in catalysis and does not need to be a wobble pair. Published by Cold Spring Harbor Laboratory Press.

Original languageEnglish (US)
Pages (from-to)1746-1760
Number of pages15
Issue number9
StatePublished - Sep 2008

All Science Journal Classification (ASJC) codes

  • Molecular Biology


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