Super-precise CRISPR tool enhanced by enzyme engineering

A super-precise CRISPR tool called base editing just got better. The group of David Liu - the inventor of base editing - has improved the accuracy of the technique by engineering enzymes that can precisely target DNA without introducing as many unwanted mutations.

Base editing, first described in 2016, has swiftly gained favour among researchers, since it offers greater control than does conventional CRISPR–Cas9 editing. On the other hand, it can introduce random, ‘off-target’ changes to the genome. The new enzymes created by the Liu group are less likely to make these errors and could allow researchers to develop safer gene therapies.

Base editing uses a so-called "nickase" - a Cas9 enzyme whose ability to cut DNA is disabled. The nickase, in turn, is fused to  enzymes that can chemically convert one DNA base to another. The function of the Cas9 nickase is to direct the fusion enzyme to the target location where the base editor will rewrite a specific letter. A potential therapeutic application of base editors is the treatment of diseases by correcting a DNA base mutation in the genome, such as sickle-cell anaemia.

Despite its popularity the technique still needs optimization. For example, the original base editors used to change the base C to T act not only on the designated target gene, but also at other random locations in the genome. Such off-target effects are concerning and may prevent application of the technique for gene therapiy in patients.

To find improved enzyme variants, Liu and colleagues inserted the base editors into bacteria and screened for resistance to an antibiotic drug. Using this method to test both naturally occurring and engineered enzymes,they identified a collection of enzymes that can convert C to T without causing as many off-target mutations as the original enzyme.

The improvements to base editing come just a few months after Liu's group described another  CRISPR technique called prime editing, which offers researchers more control over a variety of different changes to the genome.

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Anzalone, A. V. et al.Naturehttps://doi.org/10.1038/s41586-019-1711-4 (2019).