Characterization of a catalytically efficient acidic RNA-cleaving deoxyribozyme

Nucleic Acids Res. 2006 Jan 3;33(22):7164-75. doi: 10.1093/nar/gki1013. Print 2005.

Abstract

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 k(obs) of approximately 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 approximately 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 approximately 10(6)-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.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Base Sequence
  • Catalysis
  • Catalytic Domain
  • DNA, Catalytic / chemistry*
  • DNA, Catalytic / genetics
  • DNA, Catalytic / metabolism
  • Genetic Engineering
  • Hydrogen-Ion Concentration
  • Kinetics
  • Molecular Sequence Data
  • Mutation
  • Nucleotides / chemistry
  • RNA / chemistry
  • RNA / metabolism*
  • Sequence Analysis, DNA
  • Sequence Deletion
  • Sulfuric Acid Esters / chemistry

Substances

  • DNA, Catalytic
  • Nucleotides
  • Sulfuric Acid Esters
  • deoxyribozyme pH4DZ1
  • RNA
  • dimethyl sulfate