Abstract
Ribozymes enhance chemical reaction rates using many of the same catalytic strategies as protein enzymes. In the hepatitis delta virus (HDV) ribozyme, site-specific self-cleavage of the viral RNA phosphodiester backbone1,2,3 requires both divalent cations and a cytidine nucleotide4,5,6. General acid–base catalysis7,8,9,10,11,12, substrate destabilization1,13 and global and local conformational changes14,15 have all been proposed to contribute to the ribozyme catalytic mechanism. Here we report ten crystal structures of the HDV ribozyme in its pre-cleaved state, showing that cytidine is positioned to activate the 2′-OH nucleophile in the precursor structure. This observation supports its proposed role as a general base in the reaction mechanism. Comparison of crystal structures of the ribozyme in the pre- and post-cleavage states reveals a significant conformational change in the RNA after cleavage and that a catalytically critical divalent metal ion from the active site is ejected. The HDV ribozyme has remarkable chemical similarity to protein ribonucleases and to zymogens for which conformational dynamics are integral to biological activity. This finding implies that RNA structural rearrangements control the reactivity of ribozymes and ribonucleoprotein enzymes.
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Acknowledgements
Diffraction data for this study were measured with the guidance of C. Ralston, J. Holton and co-workers. We thank M. Been, K. Karbstein, J. Piccirilli, R. Spanggord and N. Walter for comments on the manuscript and for sharing unpublished data. This work was supported in part by the NIH.
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Supplementary information
Supplementary Table S1
Crystallography statistics for ten structures of the wild type and C75U mutant precursor forms of the hepatitis delta virus (HDV) ribozyme. (DOC 61 kb)
Supplementary Figure S1
Identification of a metal ion in the wild type precursor ribozyme active site. (PPT 428 kb)
Supplementary Figure S2
Competitive inhibition of C75U mutant ribozyme reactivity by cobalt hexammine. (PPT 28 kb)
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Ke, A., Zhou, K., Ding, F. et al. A conformational switch controls hepatitis delta virus ribozyme catalysis. Nature 429, 201–205 (2004). https://doi.org/10.1038/nature02522
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DOI: https://doi.org/10.1038/nature02522
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