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Nucleic Acids Research 2006 33(22):7164-7175; doi:10.1093/nar/gki1013
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Published online 3 January 2006

© The Author 2005. Published by Oxford University Press. All rights reserved
The online version of this article has been published under an open access model. Users are entitled to use, reproduce, disseminate, or display the open access version of this article for non-commercial purposes provided that: the original authorship is properly and fully attributed; the Journal and Oxford University Press are attributed as the original place of publication with the correct citation details given; if an article is subsequently reproduced or disseminated not in its entirety but only in part or as a derivative work this must be clearly indicated. For commercial re-use, please contact journals.permissions{at}oxfordjournals.org

Article

Characterization of a catalytically efficient acidic RNA-cleaving deoxyribozyme

Srinivas A. Kandadai and Yingfu Li*

Department of Biochemistry and Biomedical Sciences and Department of Chemistry, McMaster University 1200 Main Street West, Hamilton, Ontario, Canada L8N 3Z5

*To whom correspondence should be addressed. Tel: +1 905 525 9140; Fax: +1 905 522 9033; Email: liying{at}mcmaster.ca

Received August 23, 2005. Revised November 11, 2005. Accepted November 25, 2005.

We previously demonstrated—through the isolation of RNA-cleaving deoxyribozymes by in vitro selection that are catalytically active in highly acidic solutions—that DNA, despite its chemical simplicity, could perform catalysis under challenging chemical conditions [Liu,Z., Mei,S.H., Brennan,J.D. and Li,Y. (2003) J. Am. Chem. Soc. 125, 7539–7545]. One remarkable DNA molecule therefrom is pH4DZ1, a self-cleaving deoxyribozyme that exhibits a kobs of ~1 min–1 at pH 3.8. In this study, we carried out a series of experiments to examine the sequence and catalytic properties of this acidic deoxyribozyme. Extensive nucleotide truncation experiments indicated that pH4DZ1 was a considerably large deoxyribozyme, requiring ~80 out of the original 123 nt for the optimal catalytic activity. A reselection experiment identified ten absolutely conserved nucleotides that are distributed in three catalytically crucial sequence elements. In addition, a trans deoxyribozyme was successfully designed. Comparison of the observed rate constant of pH4DZ1 with experimentally determined rate constant for the uncatalyzed reaction revealed that pH4DZ1 achieved a rate enhancement of ~106-fold. This study provides valuable information about this low-pH-functional deoxyribozyme and paves way for further structural and mechanistic characterization of this unique catalytic DNA.


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