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Related Experiment Videos

Directed evolution of nucleic acid enzymes.

Gerald F Joyce1

  • 1Department of Chemistry and The Skaggs Institute for Chemical Biology, The Scripps Research Institute, La Jolla, California 92037, USA. gjoyce@scripps.edu

Annual Review of Biochemistry
|June 11, 2004
PubMed
Summary

In vitro evolution, a lab technique mimicking Darwinian evolution, creates novel nucleic acid enzymes. This review examines RNA enzymes for ligation and DNA enzymes for cleavage, highlighting their catalytic mechanisms and complexities.

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Area of Science:

  • Biochemistry
  • Molecular Biology
  • Synthetic Biology

Background:

  • Darwinian evolution in nature yields sophisticated enzymes.
  • In vitro evolution is a powerful laboratory method for enzyme development.
  • Nucleic acid enzymes (ribozymes and DNAzymes) offer catalytic functions.

Purpose of the Study:

  • To review the principles and practices of in vitro evolution for creating nucleic acid enzymes.
  • To illustrate the development of RNA enzymes for template-directed ligation and DNA enzymes for RNA cleavage.
  • To discuss the catalytic mechanisms and complexities of these in vitro evolved enzymes.

Main Methods:

  • In vitro evolution of random-sequence RNA and DNA libraries.
  • Selection and amplification of catalytic nucleic acid variants.

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  • Characterization of enzymatic activity and catalytic mechanisms.

    • Main Results:

    • Demonstration of RNA enzymes catalyzing template-directed RNA ligation.
    • Development of DNA enzymes capable of RNA cleavage.
    • Analysis of reaction mechanisms, including cofactor dependence for DNA enzymes.

    Conclusions:

    • In vitro evolution is a versatile strategy for generating novel nucleic acid catalysts.
    • The complexity of evolved enzymes correlates with reaction difficulty and cofactor requirements.
    • Understanding these catalytic nucleic acids advances molecular biology and synthetic chemistry.