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Re-engineering the adenine deaminase TadA-8e for efficient and specific CRISPR-based cytosine base editing
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Re-engineering the adenine deaminase TadA-8e for efficient and specific CRISPR-based cytosine base editing
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Journal Article
Re-engineering the adenine deaminase TadA-8e for efficient and specific CRISPR-based cytosine base editing
2023
Overview
Cytosine base editors (CBEs) efficiently generate precise C·G-to-T·A base conversions, but the activation-induced cytidine deaminase/apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like (AID/APOBEC) protein family deaminase component induces considerable off-target effects and indels. To explore unnatural cytosine deaminases, we repurpose the adenine deaminase TadA-8e for cytosine conversion. The introduction of an N46L variant in TadA-8e eliminates its adenine deaminase activity and results in a TadA-8e-derived C-to-G base editor (Td-CGBE) capable of highly efficient and precise C·G-to-G·C editing. Through fusion with uracil glycosylase inhibitors and further introduction of additional variants, a series of Td-CBEs was obtained either with a high activity similar to that of BE4max or with higher precision compared to other reported accurate CBEs. Td-CGBE/Td-CBEs show very low indel effects and a background level of Cas9-dependent or Cas9-independent DNA/RNA off-target editing. Moreover, Td-CGBE/Td-CBEs are more efficient in generating accurate edits in homopolymeric cytosine sites in cells or mouse embryos, suggesting their accuracy and safety for gene therapy and other applications.
Improved cytosine base editors are created by repurposing an adenine deaminase.
Publisher
Nature Publishing Group US,Nature Publishing Group
Subject
/ Activation-induced cytidine deaminase
/ Adenine
/ Aminohydrolases - metabolism
/ Animals
/ Biology
/ Biomedical and Life Sciences
/ Biomedical Engineering/Biotechnology
/ CRISPR
/ CRISPR-Cas Systems - genetics
/ Cytidine Deaminase - genetics
/ Cytidine Deaminase - metabolism
/ Cytosine
/ Editing
/ Editors
/ Embryos
/ Enzymes
/ Genomes
/ Mice
/ mRNA
/ Proteins
/ RNA
/ Uracil
