Cytosine Base Editor C-terminal Fragment

C-terminal fragment of cytosine base editor split via Npu intein at C574. Pairs with N-terminal fragment for editor reconstitution.

Length: 1000 bp (333 aa)

Type: Cytosine Base Editor

Conversion: C-to-T

Origin: SpCas9 D10A nickase (amino acids 574-1368) with Npu intein C-terminal domain

Characteristics

Contains C-terminal portion of SpCas9 D10A (residues 574-1368) including PAM recognition and DNA binding domains. Split site at C574 enables Npu intein trans-splicing with N-terminal fragment. Reference AAV construct includes U6-sgRNA expression cassette with BsmBI cloning sites for protospacer insertion. Cbh promoter drives Cas9 fragment expression.

Applications

Dual-delivery cytosine base editing systems requiring split architecture. Includes sgRNA expression for single-vector guide RNA delivery in C-terminal construct. Validated for AAV-mediated delivery across multiple tissues. Can be adapted to other delivery platforms when combined with N-terminal fragment.

Limitations

Non-functional without N-terminal pair. Total coding sequence of reconstituted editor approaches AAV packaging limits. Requires BsmBI digestion and ligation to insert target-specific protospacer sequences in reference construct. Must be co-delivered with split-cbe-nt for C-to-T editing activity.

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. 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