UMMS Affiliation

RNA Therapeutics Institute

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Bioinformatics | Computational Biology | Enzymes and Coenzymes | Genetic Phenomena | Genetics and Genomics | Nucleic Acids, Nucleotides, and Nucleosides


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[Formula: see text]-NAAN-3[Formula: see text] 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.


CRISPR-Cas9 genome editing, Computational biology and bioinformatics, Synthetic biology

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© The Author(s) 2020. Open Access: This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit

DOI of Published Version



Chatterjee P, Lee J, Nip L, Koseki SRT, Tysinger E, Sontheimer EJ, Jacobson JM, Jakimo N. A Cas9 with PAM recognition for adenine dinucleotides. Nat Commun. 2020 May 18;11(1):2474. doi: 10.1038/s41467-020-16117-8. PMID: 32424114; PMCID: PMC7235249. Link to article on publisher's site

Journal/Book/Conference Title

Nature communications

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Creative Commons License

Creative Commons Attribution 4.0 License
This work is licensed under a Creative Commons Attribution 4.0 License.