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[Report] Structures of a CRISPR-Cas9 R-loop complex primed for DNA cleavage

ScienceNOW - Fetched: February 18th, 2016, 3:00pm UTC

Bacterial adaptive immunity and genome engineering involving the CRISPR (clustered regularly interspaced short palindromic repeats)–associated (Cas) protein Cas9 begin with RNA-guided DNA unwinding to form an RNA-DNA hybrid and a displaced DNA strand inside the protein. The role of this R-loop structure in positioning each DNA strand for cleavage by the two Cas9 nuclease domains is unknown. We determine molecular structures of the catalytically active Streptococcus pyogenes Cas9 R-loop that show the displaced DNA strand located near the RuvC nuclease domain active site. These protein-DNA interactions, in turn, position the HNH nuclease domain adjacent to the target DNA strand cleavage site in a conformation essential for concerted DNA cutting. Cas9 bends the DNA helix by 30°, providing the structural distortion needed for R-loop formation. Authors: Fuguo Jiang, David W. Taylor, Janice S. Chen, Jack E. Kornfeld, Kaihong Zhou, Aubri J. Thompson, Eva Nogales, Jennifer A. Doudna

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