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Dissecting metal ion–dependent folding and catalysis of a single DNAzyme

Nature Chemical Biology volume 3, pages 763–768 (2007)Cite this article

Abstract

Protein metalloenzymes use various modes for functions for which metal-dependent global conformational change is required in some cases but not in others. In contrast, most ribozymes require a global folding that almost always precedes enzyme reactions. Herein we studied metal-dependent folding and cleavage activity of the 8–17 DNAzyme using single-molecule fluorescence resonance energy transfer. Addition of Zn2+ and Mg2+ induced folding of the DNAzyme into a more compact structure followed by a cleavage reaction, which suggests that the DNAzyme may require metal-dependent global folding for activation. In the presence of Pb2+, however, the cleavage reaction occurred without a precedent folding step, which suggests that the DNAzyme may be prearranged to accept Pb2+ for the activity. Neither ligation reaction of the cleaved substrates nor dynamic changes between folded and unfolded states was observed. These features may contribute to the unusually fast Pb2+-dependent reaction of the DNAzyme. These results suggest that DNAzymes can use all modes of activation that metalloproteins use.

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Figure 1: The proposed secondary structure of the 8–17 DNAzyme and its fluorophore attachments for the smFRET studies.
Figure 2: Zn2+-dependent conformational changes and cleavage reaction.
Figure 3: Mg2+-dependent conformational changes and cleavage reaction.
Figure 4: Pb2+-dependent conformational changes and cleavage reaction.

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Acknowledgements

We thank C. Joo and R. Roy for generous help with performing experiments and data analysis, and J.H. Lee for help with drawing the graphical abstract. This material is based on work supported by the US Department of Energy (DEFG02–01–ER63179), the US National Institutes of Health (GM065367) and the US National Science Foundation (CTS–0120978 and DMI- 0328162). T.H. is a Howard Hughes Medical Institute Investigator.

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  1. Ivan Rasnik

    Present address: Present address: Emory University, Department of Physics, 400 Dowman Drive, Atlanta, Georgia 30322-2430, USA.,

Authors and Affiliations

  1. Department of Chemistry, University of Illinois at Urbana-Champaign, 600 South Mathews Avenue, Urbana, 61801, Illinois, USA

    Hee-Kyung Kim, Juewen Liu & Yi Lu

  2. Department of Physics, University of Illinois at Urbana-Champaign, 1110 West Green Street, Urbana, 61801-3080, Illinois, USA

    Ivan Rasnik & Taekjip Ha

  3. Howard Hughes Medical Institute, 1110 West Green Street, Urbana, 61801, Illinois, USA

    Taekjip Ha

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  1. Hee-Kyung Kim
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Contributions

H.-K.K. designed and carried out all the experiments and wrote the manuscript; I.R. designed and carried out smFRET experiments; J.L. designed smFRET experiments; T.H. designed smFRET experiments and wrote the manuscript; Y.L. designed all the experiments and wrote the manuscript.

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Correspondence to Taekjip Ha or Yi Lu.

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Kim, HK., Rasnik, I., Liu, J. et al. Dissecting metal ion–dependent folding and catalysis of a single DNAzyme. Nat Chem Biol 3, 763–768 (2007). https://doi.org/10.1038/nchembio.2007.45

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