Synthesis of programmable integrases.
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.
Gordley
Russell M
RM
Departments of Molecular Biology and Chemistry and The Skaggs Institute for Chemical Biology, BCC 550, The Scripps Research Institute, La Jolla, CA 92037, USA.
Gersbach
Charles A
CA
Barbas
Carlos F
CF
D001665:Binding Sites; D020134:Catalytic Domain; D004268:DNA-Binding Proteins / Q000138:chemical synthesis / Q000235:genetics; D055879:Gene Knock-In Techniques / Q000379:methods; D018390:Gene Targeting / Q000379:methods*; D015894:Genome, Human; D006801:Humans; D019426:Integrases / Q000138:chemical synthesis* / Q000235:genetics; D015202:Protein Engineering / Q000379:methods; D011993:Recombinant Fusion Proteins / Q000138:chemical synthesis* / Q000235:genetics; D045522:Recombinases / Q000138:chemical synthesis / Q000235:genetics; D013379:Substrate Specificity; D019076:Transgenes; D016335:Zinc Fingers
D004268:DNA-Binding Proteins; D011993:Recombinant Fusion Proteins; D045522:Recombinases; D019426:Integrases