Synthesis of programmable integrases

Proc Natl Acad Sci U S A. 2009 Mar 31;106(13):5053-8. doi: 10.1073/pnas.0812502106. Epub 2009 Mar 12.

Abstract

Accurate modification of the 3 billion-base-pair human genome requires tools with exceptional sequence specificity. Here, we describe a general strategy for the design of enzymes that target a single site within the genome. We generated chimeric zinc finger recombinases with cooperative DNA-binding and catalytic specificities that integrate transgenes with >98% accuracy into the human genome. These modular recombinases can be reprogrammed: New combinations of zinc finger domains and serine recombinase catalytic domains generate novel enzymes with distinct substrate sequence specificities. Because of their accuracy and versatility, the recombinases/integrases reported in this work are suitable for a wide variety of applications in biological research, medicine, and biotechnology where accurate delivery of DNA is desired.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Binding Sites
  • Catalytic Domain
  • DNA-Binding Proteins / chemical synthesis
  • DNA-Binding Proteins / genetics
  • Gene Knock-In Techniques / methods
  • Gene Targeting / methods*
  • Genome, Human*
  • Humans
  • Integrases / chemical synthesis*
  • Integrases / genetics
  • Protein Engineering / methods
  • Recombinant Fusion Proteins / chemical synthesis*
  • Recombinant Fusion Proteins / genetics
  • Recombinases / chemical synthesis
  • Recombinases / genetics
  • Substrate Specificity
  • Transgenes
  • Zinc Fingers

Substances

  • DNA-Binding Proteins
  • Recombinant Fusion Proteins
  • Recombinases
  • Integrases