High-efficiency base editing for nuclear and mitochondrial DNA with an optimized DYW-like deaminase

CRISPR-based cytosine base editors enable precise genome editing without inducing double-stranded DNA breaks yet traditionally depend on a limited selection of deaminases from the APOBEC/AID or TadA families. Here, we present SsCBE, a CRISPR-based cytosine base editor utilizing SsdA_tox, a DYW-like deaminase derived from the toxin of Pseudomonas syringae. Strategic engineering of SsdA_tox has led to remarkable improvements in the base editing efficiency (by up to 8.4-fold) and specificity for SsCBE, while concurrently reducing cytotoxicity. Exhibiting exceptional versatility, SsCBE was delivered and efficiently applied using diverse delivery methods, including engineered virus-like particles. Its application has enabled targeted cytosine base editing in mouse zygotes and pioneering edits in mitochondrial DNA. SsCBE expands the genome editing toolbox by introducing a distinct deaminase scaffold with broad utility for both basic research and potential therapeutic applications.