Reprogramming homing endonuclease specificity through computational design and directed evolution
By Summer B. Thyme, Sandrine J. S. Boissel, S. Quadri, T. Nolan, David Baker, R. Park, Lara Kusak, Justin Ashworth. 2013
39 citations recorded by the source corpus.
Homing endonucleases (HEs) can be used to induce targeted genome modification to reduce the fitness of pathogen vectors such as the malaria-transmitting Anopheles gambiae and to correct deleterious mutations in genetic diseases. We describe the creation of an extensive set of HE variants with novel DNA cleavage specificities using an integrated experimental and computational approach. Using computational modeling and an improved selection strategy, which optimizes specificity in addition to activity, we engineered an endonuclease to cleave in a gene associated with Anopheles sterility and another to cleave near a mutation that causes pyruvate kinase deficiency. In the course of this work we observed unanticipated context-dependence between bases which will need to be mechanistically understood for reprogramming of specificity to succeed more generally.
Subfields: Molecular Biology
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In Molecular Biology
- David Baker
- Philip Bradley
- Yifan Song
- Summer B. Thyme
- Lara Kusak
- George M. Church
- Tanja Kortemme
- T. Brunette
Citations in the corpus
- Massively parallel determination and modeling of endonuclease substrate specificity (2014, methodology): “tivity by linking bacterial survival to target-site cleavage (14,42).”
- Concerning RNA-guided gene drives for the alteration of wild populations (2014, background): “Numerous laboratories have sought to accomplish this goal for well over a decade with only a few recent successes (Chan et al., 2013a; Thyme et al., 2013; Takeuchi et al., 2014).”