New Cas9 protein: SaCas9-HF

Our development of this new SaCas9 provides an alternative to the wild-type Cas9 toolbox, where highly precise genome editing is needed.

Zheng Zongli

SaCas9-HF – after Staphylococcus aureus CRISPR-associated protein 9 with high fidelity – was engineered by scientists from Hong Kong (CityU and Karolinska collaboration). This is the second high fidelity SaCas9, following eSaCas9 and developments around more popular SpCas9, which already has a few designed variants.

SaCas9 differs from SpCas9 in recognized motif – 5′-NNGRRT-3′ – and size – SaCas9 has over 300 amino acid less, which corresponds to over 900 nucleotides less in plasmids. The latter is especially useful in viral therapies, where the most popular adeno-associated virus (AAV) can carry up to ~5 thousand nucleotides.

Researchers modified four amino acids close to DNA-contact region. Created SaCas9-HF was shown to induce less off-target mutations in experiments with GUIDE-seq and with deep sequencing. At the same time, normal (on-target) activity of Cas9 was lower – in epithelial cells and with AAV delivery, SaCas9-HF modified 18% cells, in comparison to 51% with wild-type Sa-Cas9.

Publication: Tan, Y., Chu, A. H. Y., Bao, S., Hoang, D. A., Kebede, F. T., Xiong, W., … Zheng, Z. (2019). Rationally engineered Staphylococcus aureus Cas9 nucleases with high genome-wide specificity. Proceedings of the National Academy of Sciences, 201906843. doi:10.1073/pnas.1906843116

Leave a Reply