Mapping the sugar dependency for rational generation of a DNA-RNA hybrid-guided Cas9 endonuclease

Fernando Orden Rueda, Michal Bista, Matthew D. Newton, Anne U. Goeppert, M. Emanuela Cuomo, Euan Gordon, Felix Kröner, Jon A. Read, Jonathan D. Wrigley, David Rueda and Benjamin J. M. Taylor ()
Additional contact information
Fernando Orden Rueda: AstraZeneca
Michal Bista: AstraZeneca
Matthew D. Newton: Imperial College London
Anne U. Goeppert: AstraZeneca
M. Emanuela Cuomo: AstraZeneca
Euan Gordon: AstraZeneca
Felix Kröner: Dynamic Biosensors GmbH
Jon A. Read: AstraZeneca
Jonathan D. Wrigley: AstraZeneca
David Rueda: Imperial College London
Benjamin J. M. Taylor: AstraZeneca

Nature Communications, 2017, vol. 8, issue 1, 1-11

Abstract: Abstract The CRISPR–Cas9 RNA-guided endonuclease system allows precise and efficient modification of complex genomes and is continuously developed to enhance specificity, alter targeting and add new functional moieties. However, one area yet to be explored is the base chemistry of the associated RNA molecules. Here we show the design and optimisation of hybrid DNA–RNA CRISPR and tracr molecules based on structure-guided approaches. Through careful mapping of the ribose requirements of Cas9, we develop hybrid versions possessing minimal RNA residues, which are sufficient to direct specific nuclease activity in vitro and in vivo with reduced off-target activity. We identify critical regions within these molecules that require ribose nucleotides and show a direct correlation between binding affinity/stability and cellular activity. This is the first demonstration of a non-RNA-guided Cas9 endonuclease and first step towards eliminating the ribose dependency of Cas9 to develop a XNA-programmable endonuclease.

Date: 2017
References: Add references at CitEc
Citations: View citations in EconPapers (3)

Downloads: (external link)
https://www.nature.com/articles/s41467-017-01732-9 Abstract (text/html)

Related works:
This item may be available elsewhere in EconPapers: Search for items with the same title.

Export reference: BibTeX RIS (EndNote, ProCite, RefMan) HTML/Text

Persistent link: https://EconPapers.repec.org/RePEc:nat:natcom:v:8:y:2017:i:1:d:10.1038_s41467-017-01732-9

Ordering information: This journal article can be ordered from
https://www.nature.com/ncomms/

DOI: 10.1038/s41467-017-01732-9

Access Statistics for this article

Nature Communications is currently edited by Nathalie Le Bot, Enda Bergin and Fiona Gillespie

More articles in Nature Communications from Nature
Bibliographic data for series maintained by Sonal Shukla () and Springer Nature Abstracting and Indexing ().