The chemistry of Cas9 and its CRISPR colleagues

RNA-guided binding and cleavage of nucleic acids by CRISPR-Cas systems is a defining feature of bacterial and archaeal adaptive immunity against viruses and plasmids. As a result of their programmable ability to cut specific DNA and RNA sequences, Cas9 and related single-subunit effector proteins from CRISPR-Cas systems have been widely adopted for research and therapeutic genome engineering applications. In this Review, we discuss the chemistry of macromolecules involved in the multistep interference pathway used by CRISPR-Cas systems that mediate accurate nucleic acid target recognition and cutting. Although this Review mainly focuses on DNA interference by Cas9, we briefly explore nucleic acid targeting by the single-effector proteins Cas12 and Cas13 to emphasize the conserved themes of precision DNA and RNA cleavage within class 2 CRISPR-Cas systems. We further highlight the unique mechanisms of surveillance complex formation, substrate recognition and target cleavage in molecular detail across diverse single-subunit CRISPR-Cas interference proteins.