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DNA capture from an adenine-based editor guided by CRISPR-Cas9



Secrets of a fast editor

CRISPR-Cas9 base editors contain RNA-controlled Cas proteins condensed with an enzyme that can deaminate DNA nucleoside. No natural enzyme deaminated adenine in DNA, leading to a breakthrough when natural transfer RNA deaminase was fused to Cas9 and evolved to give an adenine-based editor (ABE) that works on DNA. Further evolution was provided by the enzyme ABE8e, which catalyzes deamination more than 1000 times faster than early ABE. Lapinaite and others. there is now a DNA-related 3.2-angstrom ABE8e resolution structure in which the target adenine is replaced by an analog designed to capture the catalytic conformation. The structure, together with kinetic data comparing ABE8e with earlier ABEs, explains how ABE8e edits DNA databases and could inform the future design of a basic editor.

science, this issue page 566

abstract

CRISPR-Cas-oriented main editors convert A • T to G • C or C • G to T • A into cellular DNA for precise genome editing. To understand the molecular basis for adenosine DNA deamination by adenine-based editors (ABEs), we determined the cryo-electron microscopic structure of ABE8e with a 3.2-anstrima resolution in a substrate-bound state in which the deaminase domain engages DNA exposed in CRISPR-Cas9 R-cycle complex. Kinetic and structural data suggest that ABE8e catalyzes deamination of DNA up to 1

1100 times faster than earlier ABEs due to mutations that stabilize DNA substrates in a limited, transfer RNA-like conformation. In addition, the accelerated DNA deamination of ABE8e suggests a previously unnoticed transient melting of DNA, which may occur during double-stranded DNA monitoring by CRISPR-Cas9. These results explain the ABE8e-mediated results of database editing and inform the future design of database editors.


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