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

• Autores: Fuguo Jiang, David W. Taylor, Janice S. Chen, Jack E. Kornfeld
• Localización: Science, ISSN 0036-8075, Vol. 351, Nº 6275, 2016, págs. 867-871
• Idioma: inglés
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• Resumen

  • 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.