Spiegelzymes® mirror-image hammerhead ribozymes and mirror-image DNAzymes, an alternative to siRNAs and microRNAs to cleave mRNAs in vivo?

PLoS One. 2014 Jan 29;9(1):e86673. doi: 10.1371/journal.pone.0086673. eCollection 2014.

Abstract

With the discovery of small non-coding RNA (ncRNA) molecules as regulators for cellular processes, it became intriguing to develop technologies by which these regulators can be applied in molecular biology and molecular medicine. The application of ncRNAs has significantly increased our knowledge about the regulation and functions of a number of proteins in the cell. It is surprising that similar successes in applying these small ncRNAs in biotechnology and molecular medicine have so far been very limited. The reasons for these observations may lie in the high complexity in which these RNA regulators function in the cells and problems with their delivery, stability and specificity. Recently, we have described mirror-image hammerhead ribozymes and DNAzymes (Spiegelzymes®) which can sequence-specifically hydrolyse mirror-image nucleic acids, such as our mirror-image aptamers (Spiegelmers) discovered earlier. In this paper, we show for the first time that Spiegelzymes are capable of recognising complementary enantiomeric substrates (D-nucleic acids), and that they efficiently hydrolyse them at submillimolar magnesium concentrations and at physiologically relevant conditions. The Spiegelzymes are very stable in human sera, and do not require any protein factors for their function. They have the additional advantages of being non-toxic and non-immunogenic. The Spiegelzymes can be used for RNA silencing and also as therapeutic and diagnostic tools in medicine. We performed extensive three-dimensional molecular modelling experiments with mirror-image hammerhead ribozymes and DNAzymes interacting with D-RNA targets. We propose a model in which L/D-double helix structures can be formed by natural Watson-Crick base pairs, but where the nucleosides of one of the two strands will occur in an anticlinal conformation. Interestingly enough, the duplexes (L-RNA/D-RNA and L-DNA/D-RNA) in these models can show either right- or left-handedness. This is a very new observation, suggesting that molecular symmetry of enantiomeric nucleic acids is broken down.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Base Pairing
  • Base Sequence
  • COS Cells
  • Chlorocebus aethiops
  • DNA, Catalytic / chemistry*
  • DNA, Catalytic / genetics
  • HeLa Cells
  • Humans
  • MicroRNAs / chemistry
  • MicroRNAs / genetics
  • Molecular Sequence Data
  • Nucleic Acid Conformation
  • RNA / chemistry*
  • RNA / genetics
  • RNA Cleavage
  • RNA Stability
  • RNA, Catalytic / chemistry*
  • RNA, Catalytic / genetics
  • RNA, Messenger / chemistry*
  • RNA, Messenger / genetics
  • RNA, Small Interfering / chemistry
  • RNA, Small Interfering / genetics
  • Stereoisomerism

Substances

  • DNA, Catalytic
  • DNA-like RNA
  • MicroRNAs
  • RNA, Catalytic
  • RNA, Messenger
  • RNA, Small Interfering
  • hammerhead ribozyme
  • RNA

Grants and funding

The authors are grateful for financial support from the Berlin RNA Network, the Federal Ministry of Science (BMBF), Deutsches Zentrum für Luft- und Raumfahrt (DLR), The Deutsche Forschungsgemeinschaft for funding of the SFB Transregio 19 (project C1), the Polish Ministry of Sciences of Higher Education, and NCN. The financial support from the Foreign Office of the Free University of Berlin is also appreciated. The funders had no role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript.