S. aureus BE3.9max-KKH C-terminal Fragment

Optimized SaCas9-KKH base editor with enhanced nuclear localization and NNNRRT PAM. Combines BE3.9max improvements with broader targeting.

Length: 1000 bp (333 aa)

Type: Cytosine Base Editor

Conversion: C-to-T

Origin: Codon-optimized APOBEC1 with enhanced NLS, fused to S. aureus Cas9-KKH D10A nickase (E782K/N968K/R1015H), split at Lys534/Cys535

Characteristics

Combines BE3.9max optimizations (enhanced NLS, codon usage) with KKH PAM flexibility (NNNRRT). Higher editing efficiency than standard saBE3-KKH through improved nuclear import and expression. Dual UGI maintains product purity with <1% indel formation.

Applications

High-efficiency C-to-T editing via dual-AAV with expanded PAM flexibility. Particularly effective in hard-to-transduce tissues requiring compact editors with broad targeting. Successfully demonstrated in brain, liver, retina, heart, and muscle with up to 59% editing.

Limitations

Large total coding sequence (~5.6 kb across both AAVs) approaches packaging limits. Requires optimization of AAV dose and serotype for each target tissue. Split-intein reconstitution efficiency varies by cell type.

Literature References

  1. Komor et al. (2016). Programmable editing of a target base in genomic DNA without double-stranded DNA cleavage. Nature - Komor 2016 BE3
  2. Komor et al. (2017). Improved base excision repair inhibition and bacteriophage Mu Gam protein yields C:G-to-T:A base editors with higher efficiency and product purity. Sci Adv - Komor 2017 BE4
  3. Koblan et al. (2018). Improving cytidine and adenine base editors by expression optimization and ancestral reconstruction. Nat Biotechnol - Koblan 2018 BE4max
  4. Levy et al. (2020). Cytosine and adenine base editing of the brain, liver, retina, heart and skeletal muscle of mice via adeno-associated viruses. Nat Biomed Eng - Levy 2020 BE3.9max