A Cas9 with PAM recognition for adenine dinucleotides

Pranam Chatterjee (), Jooyoung Lee, Lisa Nip, Sabrina R. T. Koseki, Emma Tysinger, Erik J. Sontheimer, Joseph M. Jacobson and Noah Jakimo
Additional contact information
Pranam Chatterjee: Center for Bits and Atoms
Jooyoung Lee: University of Massachusetts Medical School
Lisa Nip: Center for Bits and Atoms
Sabrina R. T. Koseki: Center for Bits and Atoms
Emma Tysinger: Center for Bits and Atoms
Erik J. Sontheimer: University of Massachusetts Medical School
Joseph M. Jacobson: Center for Bits and Atoms
Noah Jakimo: Center for Bits and Atoms

Nature Communications, 2020, vol. 11, issue 1, 1-6

Abstract: 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 II-A CRISPR-associated enzyme through identification of the natural 5$$^{\prime}$$′-NAAN-3$$^{\prime}$$′ PAM preference of Streptococcus macacae Cas9 (SmacCas9). To achieve efficient editing activity, we graft the PAM-interacting domain of SmacCas9 to its well-established ortholog from Streptococcus pyogenes (SpyCas9), and further engineer an increased efficiency variant (iSpyMac) for robust genome editing activity. We establish that our hybrids can target all adenine dinucleotide PAM sequences and possess robust and accurate editing capabilities in human cells.

Date: 2020
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DOI: 10.1038/s41467-020-16117-8

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