Probing accessible sites for ribozymes on human acetylcholinesterase RNA

RNA. 1997 Apr;3(4):429-37.


In order to design ribozymes for the efficient cleavage of a human acetylcholinesterase (AChE) in vitro transcript, a completely randomized decadeoxyribonucleotide (dN10) was used in conjunction with RNase H to identify suitable sites for annealing. Based on the observed cleavage pattern, ribozymes were designed to cleave the transcript at these positions. Five ribozymes so designed proved to be efficient in the transcript cleavage (k(react)/Km ranged from 0.9 x 10(4) to 68.2 x 10(4) M(-1) min(-10)). The best was 150-fold more active than the best designed on the basis of the MFold program. Thus, the RNase H mapping demonstrated a high predictive power for favorable ribozyme cleavage sites. The digestion pattern with RNase H differed dramatically from that observed with the single-strand-specific mung bean nuclease.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Acetylcholinesterase / genetics*
  • Humans
  • Kinetics
  • Models, Molecular
  • Nucleic Acid Conformation
  • RNA Precursors / metabolism*
  • RNA, Catalytic / metabolism*
  • RNA, Double-Stranded / metabolism
  • RNA, Messenger / metabolism*
  • Ribonuclease H / metabolism
  • Single-Strand Specific DNA and RNA Endonucleases / metabolism
  • Substrate Specificity


  • RNA Precursors
  • RNA, Catalytic
  • RNA, Double-Stranded
  • RNA, Messenger
  • Acetylcholinesterase
  • Ribonuclease H
  • Single-Strand Specific DNA and RNA Endonucleases