Journal
SCIENCE
Volume 369, Issue 6503, Pages 566-+Publisher
AMER ASSOC ADVANCEMENT SCIENCE
DOI: 10.1126/science.abb1390
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Funding
- Centers for Excellence in Genomic Science of the National Institutes of Health [RM1HG009490]
- Office of the Director, National Institutes of Health (OD) [U01AI142817-02]
- William M. Keck Foundation
- National Multiple Sclerosis Society
- Paul Allen Frontiers Group
- Howard Hughes Medical Institute
- National Science Foundation [1817593]
- NHMRC [1175568]
- American Australian Association
- Paul G. Allen Frontiers Group
- NIH [R01GM061115, T32GM113770, U01 AI142756, RM1 HG009490, R01 EB022376, R35 GM118062]
- St. Jude Collaborative Research Consortium
- Bill and Melinda Gates Foundation
- HHMI
- Harvard Chemical Biology Training Grant [T32 GM095450]
- National Health and Medical Research Council of Australia [1175568] Funding Source: NHMRC
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CRISPR-Cas-guided base editors convert A center dot T to G center dot C, or C center dot G to T center dot A, in cellular DNA for precision genome editing. To understand the molecular basis for DNA adenosine deamination by adenine base editors (ABEs), we determined a 3.2-angstrom resolution cryo-electron microscopy structure of ABE8e in a substrate-bound state in which the deaminase domain engages DNA exposed within the CRISPR-Cas9 R-loop complex. Kinetic and structural data suggest that ABE8e catalyzes DNA deamination up to similar to 1100-fold faster than earlier ABEs because of mutations that stabilize DNA substrates in a constrained, transfer RNA-like conformation. Furthermore, ABE8e's accelerated DNA deamination suggests a previously unobserved transient DNA melting that may occur during double-stranded DNA surveillance by CRISPR-Cas9. These results explain ABE8e-mediated base-editing outcomes and inform the future design of base editors.
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