Phage-assisted evolution of an adenine base editor with improved Cas domain compatibility and activity

Applications of adenine base editors (ABEs) have been constrained by the limited compatibility of the deoxyadenosine deaminase component with Cas homologs other than SpCas9. We evolved the deaminase component of ABE7.10 using phage-assisted non-continuous and continuous evolution (PANCE and PACE), resulting in ABE8e. ABE8e contains eight additional mutations that increase activity (k(app)) 590-fold compared with ABE7.10. ABE8e offers substantially improved editing efficiencies when paired with a variety of Cas9 or Cas12 homologs. ABE8e is more processive than ABE7.10, which could benefit screening, disrupting regulatory regions and multiplex base editing applications. A modest increase in Cas9-dependent and -independent DNA off-target editing, and in transcriptome-wide RNA off-target editing can be ameliorated by introducing additional mutations in the TadA-8e domain. Finally, we show that ABE8e can efficiently edit natural mutations in a GATA1 binding site in the BCL11A enhancer or the HBG promoter in human cells, targets which were poorly edited with ABE7.10. ABE8e broadens the effectiveness and applicability of adenine base editing.