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bioRxiv · 10.1101/429654

A Cas9 with Complete PAM Recognition for Adenine Dinucleotides

Jakimo, N.·Chatterjee, P.·Nip, L.·Jacobson, J. M.
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Abstract

CRISPR-associated (Cas) DNA-endonucleases are remarkably effective tools for genome engineering, but have limited target ranges due to their protospacer adjacent motif (PAM) requirements. We demonstrate a critical expansion of the targetable sequence space for a Type-IIA CRISPR-associated enzyme through identification of the natural 5-NAA-3 PAM specificity of a Streptococcus macacae Cas9 (Smac Cas9). We further recombine protein domains between Smac Cas9 and its well-established ortholog from Streptococcus pyogenes (Spy Cas9), as well as an \"increased\" nucleolytic variant (iSpy Cas9), to achieve consistent mediation of gene modification and base editing. In a comparison to previously reported Cas9 and Cas12a enzymes, we show that our hybrids recognize all adenine dinucleotide PAM sequences and possess robust editing efficiency in human cells.

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Jakimo, N., Chatterjee, P., Nip, L., Jacobson, J. M.. 2018-09-27. A Cas9 with Complete PAM Recognition for Adenine Dinucleotides. https://doi.org/10.1101/429654

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