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- [1] Thuronyi BW, 2019, NAT BIOTECHNOL, V37, P1070, DOI 10.1038/s41587-019-0193-0
Continuous evolution of base editors with expanded target compatibility and improved activity (vol 37, pg 1070, 2019)
被引：30
作者：
Thuronyi, Benjamin W.
Koblan, Luke W.
Levy, Jonathan M.
Yeh, Wei-Hsi
Zheng, Christine
Newby, Gregory A.
Wilson, Christopher
Bhaumik, Mantu
Shubina-Oleinik, Olga
Holt, Jeffrey R.
Liu, David R.
机构：
[1] Merkin Institute of Transformative Technologies in Healthcare, Broad Institute of Harvard and MIT, Cambridge, MA
[2] Department of Chemistry and Chemical Biology, Harvard University, Cambridge, MA
[3] Program in Speech and Hearing Bioscience and Technology, Harvard Medical School, Boston, MA
[4] Department of Neurology, F.M. Kirby Neurobiology Center, Boston Children’s Hospital and Harvard Medical School, Boston, MA
[5] Departments of Otolaryngology and Neurology, F.M. Kirby Neurobiology Center, Boston Children’s Hospital and Harvard Medical School, Boston, MA
[6] Howard Hughes Medical Institute, Harvard University, Cambridge, MA
[7] Department of Chemistry, Williams College, Williamstown, MA
关键词：
Efficiency;
D O I：
10.1038/s41587-019-0253-5
中图分类号：
Q81 [生物工程学（生物技术）];
Q93 [微生物学];
学科分类号：
071005 ;
0836 ;
090102 ;
100705 ;
摘要：
Base editors use DNA-modifying enzymes targeted with a catalytically impaired CRISPR protein to precisely install point mutations. Here, we develop phage-assisted continuous evolution of base editors (BE–PACE) to improve their editing efficiency and target sequence compatibility. We used BE–PACE to evolve cytosine base editors (CBEs) that overcome target sequence context constraints of canonical CBEs. One evolved CBE, evoAPOBEC1-BE4max, is up to 26-fold more efficient at editing cytosine in the GC context, a disfavored context for wild-type APOBEC1 deaminase, while maintaining efficient editing in all other sequence contexts tested. Another evolved deaminase, evoFERNY, is 29% smaller than APOBEC1 and edits efficiently in all tested sequence contexts. We also evolved a CBE based on CDA1 deaminase with much higher editing efficiency at difficult target sites. Finally, we used data from evolved CBEs to illuminate the relationship between deaminase activity, base editing efficiency, editing window width and byproduct formation. These findings establish a system for rapid evolution of base editors and inform their use and improvement. © 2019, The Author(s), under exclusive licence to Springer Nature America, Inc.
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页码：1091 / 1091
页数：1
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